Semaphorins as emerging clinical biomarkers and therapeutic targets in cancer.

Semaphorins are a large family of developmental regulatory signals, characterized by aberrant expression in human cancers. These molecules crucially control cell-cell communication, cell migration, invasion and metastasis, tumor angiogenesis, inflammatory and anti-cancer immune responses. Semaphorins comprise secreted and cell surface-exposed molecules and their receptors are mainly found in the Plexin and Neuropilin families, which are further implicated in a signaling network controlling the tumor microenvironment. Accumulating evidence indicates that semaphorins may be considered as novel clinical biomarkers for cancer, especially for the prediction of patient survival and responsiveness to therapy. Moreover, preclinical experimental studies have demonstrated that targeting semaphorin signaling can interfere with tumor growth and/or metastatic dissemination, suggesting their relevance as novel therapeutic targets in cancer; this has also prompted the development of semaphorin-interfering molecules for application in the clinic. Here we will survey, in diverse human cancers, the current knowledge about the relevance of semaphorin family members, and conceptualize potential lines of future research development in this field.

Semaphorin 4A antibody alleviates arsenic-induced hepatotoxicity in mice via inhibition of AKT2/NF-κB inflammatory signaling.

Semaphorin (Sema) 3A and Sema 4A are immunomodulatory molecules with a common receptor, neuropilin-1 (NRP-1), on the immune cells. Sema 3A binds to NRP-1 and inhibits T cell activation and inflammation, while Sema 4A binds to NRP-1 and promotes T cell activation and inflammation. These molecules are associated closely with the regulation of protein kinase B (AKT)/nuclear factor-kappaB (NF-κB) signaling, which are poorly understood in arsenic toxicity. The present study explored the role of Sema 3A or Sema 4A in arsenic-induced hepatotoxicity in mice. Arsenic exposure induced hepatic injury and resulted in the activations of p-AKT2, NF-κB p65, and NOD-like receptor family pyrin domain-containing 3 (NLRP3) inflammasome, downregulation of Sema 3A, and upregulation of Sema 4A or NRP-1. Interestingly, intervention with anti-Sema 4A antibody showed the mitigation of arsenic-induced hepatotoxicity, accompanied by the downregulation of Sema 4A, rebound of Sema 3A, and upregulation of NRP-1. And, the inflammatory signaling p-AKT2 or NF-κB p65, and NLRP3 inflammasome showed a downregulation compared with arsenic treatment group. In contrast, anti-Sema 3A antibody intervention did not show the significant effect in the histopathological features compared with arsenic treatment group. In conclusion, the anti-Sema 4A antibody antagonizes arsenic-induced hepatotoxicity in mice and may be involved in the inhibitions of AKT2/NF-κB and NLRP3 inflammatory signaling mediated synergistically by Sema 4A or Sema 3A and their receptor NRP-1.

Semaphorin-3C Is Upregulated in Polycystic Kidney Epithelial Cells and Inhibits Angiogenesis of Glomerular Endothelial Cells.

BACKGROUND: Polycystic kidney disease (PKD) is a hereditary disease characterized by cyst formation in the kidneys bilaterally. It has been observed that semaphorin-3C (SEMA3C) is overexpressed in polycystic kidney epithelial cells. It is hypothesized that upregulated SEMA3C would contribute to survival of polycystic kidney epithelial cells. Furthermore, as the kidney is a highly vascularized organ, the secreted SEMA3C from PKD epithelial cells will affect glomerular endothelial cells (GECs) in a paracrine manner. METHODS: To evaluate the effect of SEMA3C on renal cells, siSEMA3C-treated PKD epithelial cells were used for further analysis, and GECs were exposed to recombinant SEMA3C (rSEMA3C). Also, co-culture and treatment of conditioned media were employed to confirm whether PKD epithelial cells could influence on GECs via SEMA3C secretion. RESULTS: SEMA3C knockdown reduced proliferation of PKD epithelial cells. In case of GECs, exposure to rSEMA3C decreased angiogenesis, which resulted from suppressed migratory ability not cell proliferation. CONCLUSIONS: This study indicates that SEMA3C is the aggravating factor in PKD. Thus, it is proposed that targeting SEMA3C can be effective to mitigate PKD.

A pan-cancer study of class-3 semaphorins as therapeutic targets in cancer.

BACKGROUND: Initially characterized as axon guidance factors, semaphorins also have been implicated to have critical roles in multiple physiological and developmental functions, including the regulation of immune responses, angiogenesis, organ formation, and the etiology of multiple forms of cancer. Moreover, their contribution in immunity and the regulation of tumour microenvironment is becoming increasingly recognized. Here, we provide a comprehensive analysis of class-3 semaphorins, the only secreted family of genes among veterbrate semaphorins, in terms of their expression profiles and their association with patient survival. We also relate their role with immune subtypes, tumour microenvironment, and drug sensitivity using a pan-cancer study. RESULTS: Expression profiles of class-3 semaphorins (SEMA3s) and their association with patient survival and tumour microenvironment were studied in 31 cancer types using the TCGA pan-cancer data. The expression of SEMA3 family varies in different cancer types with striking inter- and intra- cancer heterogeneity. In general, our results show that SEMA3A, SEMA3C, and SEMA3F are primarily upregulated in cancer cells, while the rest of SEMA3s are mainly down-regulated in the tested tumours. The expression of SEMA3 family members was frequently associated with patient overall survival. However, the direction of the association varied with regards to the particular SEMA3 isoform queried and the specific cancer type tested. More specifically, SEMA3A and SEMA3E primarily associate with a poor prognosis of survival, while SEMA3G typically associates with survival advantage. The rest of SEMA3s show either survival advantage or disadvantage dependent on cancer type. In addition, all SEMA3 genes show significant association with immune infiltrate subtypes, and they also correlate with level of stromal cell infiltration and tumour cell stemness with various degrees. Finally, our study revealed that SEMA3 genes, especially SEMA3C and SEMA3F may contribute to drug induced cancer cell resistance. CONCLUSIONS: Our systematic analysis of class-3 semaphorin gene expression and their association with immune infiltrates, tumour microenvironment and cancer patient outcomes highlights the need to study each SEMA3 member as a separate entity within each specific cancer type. Also our study validated the identification of class-3 semaphorin signals as promising therapeutic targets in cancer although further laboratory validation still needed.

SEMA3B-AS1-inhibited osteogenic differentiation of human mesenchymal stem cells revealed by quantitative proteomics analysis.

Human mesenchymal stem cells (hMSCs) are fibroblastoid multipotent adult stem cells with capacities of differentiation into osteoblasts and chondrocytes and show great potential in new bone formation and bone repair-related clinical settings, such as osteoporosis. Long noncoding RNAs (lncRNAs) have been demonstrated to play important roles in various biological processes. Here, we report an antisense lncRNA SEMA3B-AS1 regulating hMSCs osteogenesis. SEMA3B-AS1 is proximal to a member of the semaphorin family Sema3b. Overexpression of SEMA3B-AS1 using the lentivirus system markedly inhibits the proliferation of hMSCs and meanwhile reduces osteogenic differentiation. Using a comprehensive proteomic technique named isobaric tag for relative and absolute quantitation, we found that SEMA3B-AS1 significantly alters the process of osteogenesis through downregulating the expression of proteins involved in actin cytoskeleton, focal adhesion, and extracellular matrix-receptor interaction, while increasing the expression of proteins in the spliceosome. Collectively, we find that SEMA3B-AS1 is a target for controlling osteogenesis of hMSCs.

The role of sema4D in vasculogenic mimicry formation in non-small cell lung cancer and the underlying mechanisms.

Vasculogenic mimicry (VM) is a special vascular pattern in malignant tumors, which is composed of highly aggressive tumor cells. This tumor cell-mediated blood supply pattern is closely associated with a poor prognosis in cancer patients. The interaction of axon guidance factor Sema4D and its high affinity receptor plexinB1 could activate small GTPase RhoA and its downstream ROCKs; this process has an active role in the migration of endothelial cells and tumor angiogenesis. Here, we have begun to uncover the role of this pathway in VM formation in non-small cell lung cancer (NSCLC). First, we confirmed this special form of vasculature in NSCLC tissues and found the existence of VM channels in tumor tissues was correlated with Sema4D expression. Further, we found that inhibition of Sema4D in the human NSCLC cells H1299 and HCC827 reduces VM formation both in vitro and in vivo. Moreover, we demonstrated that downregulating the expression of plexinB1 by siRNA expressing vectors and inhibiting the RhoA/ROCK signaling pathway using fasudil can reduce VM formation of H1299 and HCC827 cells. Finally, we found that suppression of Sema4D leads to less stress fibers and depleted the motility of H1299 and HCC827 cells. Collectively, our study implicates Sema4D plays an important role in the process of VM formation in NSCLC through activating the RhoA/ROCK pathway and regulating tumor cell plasticity and migration. Modulation of the Sema4D/plexinB1 and downstream RhoA/ROCK pathway may prevent the tumor blood supply through the VM pattern, which may eventually halt growth and metastasis of NSCLC.

SEMA3C drives cancer growth by transactivating multiple receptor tyrosine kinases via Plexin B1.

Growth factor receptor tyrosine kinase (RTK) pathway activation is a key mechanism for mediating cancer growth, survival, and treatment resistance. Cognate ligands play crucial roles in autocrine or paracrine stimulation of these RTK pathways. Here, we show SEMA3C drives activation of multiple RTKs including EGFR, ErbB2, and MET in a cognate ligand-independent manner via Plexin B1. SEMA3C expression levels increase in castration-resistant prostate cancer (CRPC), where it functions to promote cancer cell growth and resistance to androgen receptor pathway inhibition. SEMA3C inhibition delays CRPC and enzalutamide-resistant progression. Plexin B1 sema domain-containing:Fc fusion proteins suppress RTK signaling and cell growth and inhibit CRPC progression of LNCaP xenografts post-castration in vivo SEMA3C inhibition represents a novel therapeutic strategy for treatment of advanced prostate cancer.

Suppression of tumor-derived Semaphorin 7A and genetic ablation of host-derived Semaphorin 7A impairs tumor progression in a murine model of advanced breast carcinoma.

Solid tumors can generate a plethora of neurogenesis-related molecules that enhance their growth and metastasis. Among them, we have identified axonal guidance molecule Semaphorin 7A (SEMA7A) in breast cancer. The goal of this study was to determine the therapeutic effect of suppressing SEMA7A levels in the 4T1 murine model of advanced breast carcinoma. We used anti-SEMA7A short hairpin RNA (shRNA) to gene silence SEMA7A in 4T1 mammary tumor cells. When implanted into the mammary fat pads of syngeneic mice, SEMA7A shRNA-expressing 4T1 tumors exhibited decreased growth rates, deferred metastasis and reduced mortality. In vitro, SEMA7A shRNA-expressing 4T1 cells had weakened proliferative, migratory and invasive abilities, and decreased levels of mesenchymal factors. Atomic force microscopy studies showed that SEMA7A shRNA-expressing 4T1 cells had an increase in cell stiffness that corresponded with their decreased malignant potential. Genetic ablation of host-derived SEMA7A further enhanced the antitumor effects of SEMA7A shRNA gene silencing in 4T1 cells. Our preclinical findings demonstrate a critical role for SEMA7A in mediating mammary tumor progression.

Sprouting strategies and dead ends in anti-angiogenic targeting of NETs.

Neuroendocrine tumors (NETs) are a heterogeneous group of neoplasms that arise from cells of the neuroendocrine system. NETs are characterized by being highly vascularized tumors that produce large amounts of proangiogenic factors. Due to their complexity and heterogeneity, progress in the development of successful therapeutic approaches has been limited. For instance, standard chemotherapy-based therapies have proven to be poorly selective for tumor cells and toxic for normal tissues. Considering the urge to develop an efficient therapy to treat NET patients, vascular targeting has been proposed as a new approach to block tumor growth. This review provides an update of the mechanisms regulating different components of vessels and their contribution to tumor progression in order to develop new therapeutic drugs. Following the description of classical anti-angiogenic therapies that target VEGF pathway, new angiogenic targets such as PDGFs, EGFs, FGFs and semaphorins are further explored. Based on recent research in the field, the combination of therapies that target multiple and different components of vessel formation would be the best approach to specifically target NETs and inhibit tumor growth.

Decreased expression of semaphorin 3D is associated with genesis and development in colorectal cancer.

BACKGROUND: Semaphorin 3D (SEMA3D) plays important roles in the genesis and progress of many cancers. However, the relationship between SEMA3D and colorectal cancer (CRC) remains unknown. The aim of this study was to investigate whether SEMA3D can be used as a predictive marker for the diagnosis, metastasis, and prognosis of CRC by assessing the expression of SEMA3D in the tissues and serum of CRC patients. METHODS: Real-time quantitative polymerase chain reaction (qPCR) was used to measure the expression of SEMA3D mRNA in 100 CRC tissues and matched normal tissues. qPCR was also used to detect the expression of SEMA3D mRNA in the CRC cell line RKO. RKO cells were transfected with SEMA3D small-interring RNA (siRNA) to interfere with endogenous SEMA3D. The migratory ability of control and SEMA3D siRNA-transfected RKO cells was determined by transwell assays. Enzyme-linked immunosorbent assay (ELISA) was utilized to detect the levels of SEMA3D in the serum of 80 CRC patients and 100 normal healthy controls. The expression of SEMA3D in 215 CRC tissues was assessed using immunohistochemistry (IHC). Then, statistical analyses were adopted to assess SEMA3D protein levels and clinical pathological characteristics. RESULTS: The mRNA expression of SEMA3D was significantly lower in CRC tissues than in paired normal tissues (t = 5.027, P < 0.0001). Compared with normal healthy controls, the serum levels of SEMA3D were decreased significantly in CRC patients (t = 3.656, P = 0.0003). The expression of SEMA3D protein was linked to lymph node metastasis, and low expression led to lymph node metastasis (χ 2 = 8.415, P = 0.004). The expression of SEMA3D in CRC tissues was a favorable prognostic factor. Patients with a higher expression of SEMA3D experienced longer survival (P = 0.002, log-rank [Mantel-Cox]; Kaplan-Meier). In addition, multivariate Cox's proportional hazard model revealed that SEMA3D is an independent prognostic marker (hazard ratio [HR] 1.818, 95% CI 1.063-3.110, P = 0.029). Moreover, transwell assays showed that knocking down SEMA3D significantly increased RKO cell migration (t = 9.268, P = 0.0008). CONCLUSIONS: SEMA3D might function as a tumor suppressor during the formation and development of CRC. SEMA3D might become a predictive marker for the diagnosis, metastasis, and prognosis of CRC and provide a novel target for the prevention and treatment of CRC.

MiR-4282 suppresses proliferation and mobility of human colorectal carcinoma cells by targeting semaphorin 3E.

BACKGROUND: MicroRNAs play an important role in cancer development. Deregulation of microRNAs can lead to tumorigenesis. Class 3 semaphorin, semaphorin 3E (Sema3E), has been shown to be implicated in tumor growth and metastasis. The role of miR-4282 in regulating colorectal carcinoma and its correlation to Sema3E remain uncertain. METHODS: Real-time quantitative reverse transcription polymerase chain reaction was used to detect the levels of miR-4282 and Sema3E in colorectal carcinoma cells and colorectal tumor tissues. Sema3E protein level in cell lines and human tissues was analyzed by western blot Transient transfections of miR-4282 inhibitor or mimics were conducted to silence or overexpress miR-4282. Sema3E siRNA was transfected to knockdown Sema3E in tumor cell lines. MTT assay was employed to measure colorectal tumor cell growth. Migration and invasion of the cells were examined by trans-well assays. Luciferase reporter assays were performed to confirm miR-4282 targeted at Sema3E. RESULTS: In the present study, reduced miR-4282 expression was observed in the colorectal carcinoma cell lines and human carcinoma tissues in comparison with normal human colon cells (P<0.05) or matched non-tumor tissues (P<0.05), whereas, Sema3E was up-regulated in colorectal carcinoma cells lines (P<0.05) and human colorectal tumor tissues (P<0.05). MiR-4282 was then reduced by the inhibitor and overexpressed by its mimics transfection. It was found that miR-4282 inhibition promoted cell growth, migration and invasion (P<0.05) of HT29 and HCT116 colorectal carcinoma cells while miR-4282 overexpression suppressed cell growth and mobility (P<0.05). Sema3E was predicted as a target of miR-4282 in miRDB database. We found that miR-4282 overexpression significantly reduced luciferase activity of pRL-Sema3E-3'-UTR (P<0.05), but failed to alter the activity of pRL-sema3E-3'-UTR-mutation. Also, miR4282 overexpression suppressed Sema3E expression in the colorectal carcinoma cell lines. To further confirm the role of Sema3E suppression in the function of the colorectal carcinoma cells by miR-4282, HT29 and HCT116 cells were transfected with Sema3E siRNA. We found that cell growth, migration and invasion of HT29 and HCT116 cells were dramatically inhibited by Sema3E knockdown (P<0.05). CONCLUSIONS: Our findings suggested that miR-4282 is a tumor suppressor in colorectal carcinoma cells and exerted its inhibitory effect on the tumor cells through targeting Sema3E by inhibiting Sema3E translation or enhancing Sema3E mRNA degradation. Thus, manipulation of miR-4282 and interfere with Sema3E might represent a potential target for the treatment of colorectal cancer.

Soluble SEMA4D/CD100: A novel immunoregulator in infectious and inflammatory diseases.

SEMA4D/CD100 is a homodimeric protein belonging to the semaphorin family of axonal guidance proteins. Semaphorin family members have received increased attention lately due to their diverse functions in the immune system. SEMA4D was the first semaphorin described to have immune functions and serves important roles in T cell priming, antibody production, and cell-to-cell adhesion. Proteolytic cleavage of SEMA4D from the cell surface gives rise to a soluble fragment of SEMA4D (sSEMA4D). Similar to the transmembranal form, sSEMA4D is thought to have immunoregulatory properties. While the exact mechanisms responsible for SEMA4D shedding remain to be elucidated, emerging data have revealed associations between elevated systemic sSEMA4D levels and severity of infectious and inflammatory diseases. This review summarizes the literature concerning sSEMA4D and discusses its potential as a novel prognostic immune-biomarker and potential target for immunotherapy.

Tumor necrosis factor alpha suppresses osteogenic differentiation of MSCs by inhibiting semaphorin 3B via Wnt/ß-catenin signaling in estrogen-deficiency induced osteoporosis.

The proinflammatory cytokines, especially tumor necrosis factor alpha (TNF-α), have been shown to inhibit osteogenic differentiation of mesenchymal stem cells (MSCs) and bone formation in estrogen-deficiency-induced osteoporosis, but the mechanisms of TNF-α impaired bone formation remain poorly understood. Semaphorins have been shown to regulate cell growth, cell migration, and cell differentiation in a variety of tissues, including bone tissue. Here, we identified a novel mechanism whereby TNF-α, suppressing Semaphorin3B expression contributes to estrogen-deficiency-induced osteoporosis. In this study, we found that TNF-α could decrease Semaphorin3B expression in osteogenic differentiation of MSCs. Overexpression of Semaphorin3B in MSCs attenuated the inhibitory effects of TNF-α on MSCs proliferation and osteoblastic differentiation. Mechanistically, activation of the Wnt/ß-catenin signaling markedly rescued TNF-α-inhibited Semaphorin3B expression, suggesting that Wnt/ß-catenin signaling was involved in the regulation of Semaphorin3B expression by TNF-α. Taken together, our results revealed a novel function for Semaphorin3B and suggested that suppressed Semaphorin3B may contribute to impaired bone formation by elevated TNF-α in estrogen-deficiency-induced osteoporosis. This study may indicate a therapeutic target gene of Semaphorin3B for osteoporosis.

Generation and preclinical characterization of an antibody specific for SEMA4D.

Semaphorin 4D (SEMA4D or CD100) is a member of the semaphorin family of proteins and an important mediator of the movement and differentiation of multiple cell types, including those of the immune, vascular, and nervous systems. Blocking the binding of SEMA4D to its receptors can result in physiologic changes that may have implications in cancer, autoimmune, and neurological disease. To study the effects of blocking SEMA4D, we generated, in SEMA4D-deficient mice, a panel of SEMA4D-specific hybridomas that react with murine, primate, and human SEMA4D. Utilizing the complementarity-determining regions from one of these hybridomas (mAb 67-2), we generated VX15/2503, a humanized IgG4 monoclonal antibody that is currently in clinical development for the potential treatment of various malignancies and neurodegenerative disorders, including multiple sclerosis and Huntington's disease. This work describes the generation and characterization of VX15/2503, including in vitro functional testing, epitope mapping, and an in vivo demonstration of efficacy in an animal model of rheumatoid arthritis.

SEMA4B inhibits growth of non-small cell lung cancer in vitro and in vivo.

We have recently shown that Semaphorin 4B (SEMA4B) inhibits the invasion of non-small cell lung cancer (NSCLC) through PI3K-dependent suppression of MMP9 activation. In the current study, we evaluated whether SEMA4B may also affect the growth of NSCLC. We thus used two human NSCLC lines, A549 and Calu-3, to examine our hypothesis. We found that overexpression of SEMA4B significantly decreased NSCLC cell growth, while SEMA4B inhibition significantly increased NSCLC cell growth, both in vitro and in vivo in an implanted NSCLC model. Adaptation of SEMA4B in NSCLC cells did not alter cell apoptosis, but changed the cell proliferation. Further analyses show that SEMA4B may induce FoxO1 nuclear retention through suppressing PI3K/Akt signaling pathway, which subsequently inhibited cell growth through the direct nuclear target of FoxO1, p21. Our study thus demonstrate a role of SEMA4B in suppressing NSCLC growth, besides its role in inhibiting cell metastasis, and highlights SEMA4B as a promising therapeutic target for NSCLC.

Semaphoring mAb: a new guide in RIT in inhibiting the proliferation of human skin carcinoma.

Semaphoring is a transmembrane receptor which participates in many cytokine-mediated signal pathways that are closely related to the angiogenesis, occurrence and development of carcinoma. The present study was designed to access the effect of mono-antibody (mAb) guided radioimmunotherapy (RIT) on skin carcinoma and investigate the potential mechanisms. Semaphoring mAb was acquired from mice (Balb/c), purified with rProtein A column; purity, concentration and activity were tested with SDS-PAGE and indirect ELISA; specificity and expression on the cutanuem carcinoma line and tissue were tested by Western blotting; morphology change was assessed by microscopy. MTT assay and colony inhibition tests were carried out to test the influence on the proliferation of tumor cells; Western blotting was also carried out for expression of apoptosis-associated (caspase-3, Bax, Bcl-2) and proliferation-related (PI3K, p-Akt, Akt, p-ERK1/2, ERK1/2) proteins and analyse the change in signal pathways (PI3K/Akt and MEK/ERK). The purity of purified semaphorin mAb was 96.5% and the titer is about 1?106. Western blotting showed semaphoring mAb to have specifically binding stripes with semaphoring b1b2 protein, B16F10, and A431 cells at 39KDa, 100KDa and 130KDa, respectively. Positive expression was detected both in cutanuem carcinoma line and tissue and it mostly located in cell membranes. MMT assay revealed dose-relate and time-relate inhibitory effect of semaphorin mAb on A431 and B16F10. Colony inhibition tests also showed dose-relate inhibitory effects. Western blotting demonstrated the expression of apoptosis and proliferation-related protein and changes in signal pathway. In conclusion, we demonstrated that semaphorin is highly expressed on the tumor cell-surfaces and RIT with semaphorin mAb has effect in inhibiting proliferation and accelerating apoptosis of tumor cells.

Nonclinical Safety Evaluation of VX15/2503, a Humanized IgG4 Anti-SEMA4D Antibody.

The humanized IgG4 monoclonal antibody VX15/2503 bound with 1 to 5 nmol/L affinity to purified recombinant semaphorin 4D (SEMA4D; CD100) produced using murine, rat, cynomolgus macaque, and human sequences. The affinity for native SEMA4D expressed on macaque T lymphocytes was approximately 0.6 nmol/L. Tissues from rats and cynomolgus macaques demonstrated specific staining only with resident lymphocytes. Single-dose and one-month toxicology/PK studies used VX15/2503 dose levels of 0 to 100 mg/kg. No toxicity was observed with either species in these studies, thus the no observed adverse effect level (NOAEL) was 100 mg/kg. Cmax, exposure, and half-life values were similar for both rats and macaques. The NOAEL in a primate maximum feasible dose study was 200 mg/kg. Saturation of T-cell-associated SEMA4D occurred following administration of single doses of 0.1 mg/kg and above; five weekly injections of VX15/2503 at a dose level of 100 mg/kg produced saturation lasting for more than 120 and 130 days, respectively, for rats and primates. Macaques administered five weekly doses of VX15/2503 showed dose-dependent reductions of 2- to 3-fold in T-cell SEMA4D (cSEMA4D) expression levels compared with controls. Reduced cSEMA4D expression levels continued until serum antibody concentrations were 2 to 5 µg/mL, and thereafter normal cSEMA4D levels were restored. On the basis of these data, a phase I clinical study of the safety and tolerability of VX15/2503 was conducted, enrolling adult patients with advanced solid tumor diseases; a single-dose, dose escalation, phase I safety study was also initiated with subjects with multiple sclerosis.

Semaphorin 4D Contributes to Rheumatoid Arthritis by Inducing Inflammatory Cytokine Production: Pathogenic and Therapeutic Implications.

OBJECTIVE: Semaphorin 4D (Sema4D)/CD100 has pleiotropic roles in immune activation, angiogenesis, bone metabolism, and neural development. We undertook this study to investigate the role of Sema4D in rheumatoid arthritis (RA). METHODS: Soluble Sema4D (sSema4D) levels in serum and synovial fluid were analyzed by enzyme-linked immunosorbent assay. Cell surface expression and transcripts of Sema4D were analyzed in peripheral blood cells from RA patients, and immunohistochemical staining of Sema4D was performed in RA synovium. Generation of sSema4D was evaluated in an ADAMTS-4-treated monocytic cell line (THP-1 cells). The efficacy of anti-Sema4D antibody was evaluated in mice with collagen-induced arthritis (CIA). RESULTS: Levels of sSema4D were elevated in both serum and synovial fluid from RA patients, and disease activity markers were correlated with serum sSema4D levels. Sema4D-expressing cells also accumulated in RA synovium. Cell surface levels of Sema4D on CD3+ and CD14+ cells from RA patients were reduced, although levels of Sema4D transcripts were unchanged. In addition, ADAMTS-4 cleaved cell surface Sema4D to generate sSema4D in THP-1 cells. Soluble Sema4D induced tumor necrosis factor α (TNFα) and interleukin-6 (IL-6) production from CD14+ monocytes. IL-6 and TNFα induced ADAMTS-4 expression in synovial cells. Treatment with an anti-Sema4D antibody suppressed arthritis and reduced proinflammatory cytokine production in CIA. CONCLUSION: A positive feedback loop involving sSema4D/IL-6 and TNFα/ADAMTS-4 may contribute to the pathogenesis of RA. The inhibition of arthritis by anti-Sema4D antibody suggests that Sema4D represents a potential therapeutic target for RA.

Antibody Blockade of Semaphorin 4D Promotes Immune Infiltration into Tumor and Enhances Response to Other Immunomodulatory Therapies.

Semaphorin 4D (SEMA4D, CD100) and its receptor plexin-B1 (PLXNB1) are broadly expressed in murine and human tumors, and their expression has been shown to correlate with invasive disease in several human tumors. SEMA4D normally functions to regulate the motility and differentiation of multiple cell types, including those of the immune, vascular, and nervous systems. In the setting of cancer, SEMA4D-PLXNB1 interactions have been reported to affect vascular stabilization and transactivation of ERBB2, but effects on immune-cell trafficking in the tumor microenvironment (TME) have not been investigated. We describe a novel immunomodulatory function of SEMA4D, whereby strong expression of SEMA4D at the invasive margins of actively growing tumors influences the infiltration and distribution of leukocytes in the TME. Antibody neutralization of SEMA4D disrupts this gradient of expression, enhances recruitment of activated monocytes and lymphocytes into the tumor, and shifts the balance of cells and cytokines toward a proinflammatory and antitumor milieu within the TME. This orchestrated change in the tumor architecture was associated with durable tumor rejection in murine Colon26 and ERBB2(+) mammary carcinoma models. The immunomodulatory activity of anti-SEMA4D antibody can be enhanced by combination with other immunotherapies, including immune checkpoint inhibition and chemotherapy. Strikingly, the combination of anti-SEMA4D antibody with antibody to CTLA-4 acts synergistically to promote complete tumor rejection and survival. Inhibition of SEMA4D represents a novel mechanism and therapeutic strategy to promote functional immune infiltration into the TME and inhibit tumor progression.

Emerging role of semaphorins as major regulatory signals and potential therapeutic targets in cancer.

Semaphorins are mainly known as guidance signals in development, acting through receptors called Plexins. However, their role in cancer is rapidly emerging in the regulation of tumor angiogenesis, tumor growth, cancer cell invasiveness, and metastatic spreading. Intriguingly, activated plexins can transactivate receptor tyrosine kinases, such as MET, VEGFR2, FGFR2, and ERBB2, and lead to distinctive effects in a cell-context-dependent manner. Moreover, certain semaphorins concomitantly target endothelial and cancer cells, and can achieve remarkable inhibition of angiogenesis and tumor growth, associated with anti-metastatic activity. Altogether, these data validate the identification of semaphorin signals as promising therapeutic targets in cancer.