Intratumoral delivery of a highly active form of XCL1 enhances antitumor CTL responses through recruitment of CXCL9-expressing conventional type-1 dendritic cells.

Conventional type 1 dendritic cells (cDC1s) play a crucial role in antitumor immunity through the induction and activation of tumor-specific CD8+ cytotoxic T cells (CTLs). The chemokine XCL1 is a major chemotactic factor for cDC1s and its receptor XCR1 is selectively expressed on cDC1s. Here, we investigated the effect of intratumoral delivery of a highly active form of murine XCL1 (mXCL1-V21C/A59C) on cDC1-mediated antitumor immunity using a hydrophilic gel patch. The hydrophilic gel patch containing mXCL1-V21C/A59C increased cDC1 accumulation in the tumor masses and promoted their migration to the regional lymph nodes, resulting in enhanced induction of tumor-specific CTLs. Tumor-infiltrating cDC1s not only expressed XCR1 but also produced CXCL9, a ligand for CXCR3 which is highly expressed on CTLs and NK cells. Consequently, CTLs and NK cells were increased in the tumor masses of mice treated with mXCL1-V21C/A59C, while immunosuppressive cells such as monocyte-derived suppressive cells and regulatory T cells were decreased. We also confirmed that anti-CXCL9 treatment decreased the tumor infiltration of CTLs. The intratumoral delivery of mXCL1-V21C/A59C significantly decreased tumor growth and prolonged survival in E.G7-OVA and B16-F10 tumor-bearing mice. Furthermore, the antitumor effect of mXCL1-V21CA59C was enhanced in combination with anti-programmed cell death protein 1 treatment. Finally, using The Cancer Genome Atlas database, we found that XCL1 expression was positively correlated with tumor-infiltrating cDC1s and a better prognosis in melanoma patients. Collectively, our findings provide a novel therapeutic approach to enhance tumor-specific CTL responses through the selective recruitment of CXCL9-expressing cDC1s into the tumor masses.

Aggravation of lipopolysaccharide-induced depressive-like behavior in CCR4-deficient mice.

Increasing evidence indicates that immune abnormalities are associated with the pathogenesis of depression. CCR4 is a chemokine receptor that regulates regulatory T cell (Treg) and Th17 cell migration. Here, using a lipopolysaccharide (LPS)-induced depression mouse model, we demonstrated that CCR4 deficiency exacerbated depressive-like behavior. Tregs and M2 macrophages, but not Th17 cells, were decreased in the brain of CCR4-deficient mice. Consistently, treatment with a CCR4 inhibitor reduced Tregs and M2 macrophages in the brain and exacerbated depressive-like behavior. Thus, CCR4 may contribute to the reduction of depressive symptoms by promoting Treg recruitment to the brain and subsequent M2 macrophage polarization.

A CCR4 antagonist attenuates atopic dermatitis-like skin inflammation by inhibiting the recruitment and expansion of Th2 cells and Th17 cells.

CCR4 is a major trafficking receptor for T-helper (Th) 2 cells and Th17 cells and is considered as a potential therapeutic target for atopic dermatitis (AD). The CCR4 ligands CCL17 and CCL22 have been reported to be upregulated in the skin lesions of AD patients. Of note, thymic stromal lymphopoietin (TSLP), a master regulator of the Th2 immune response, promotes the expression of CCL17 and CCL22 in AD skin lesions. Here, we investigated the role of CCR4 in an AD mouse model induced by MC903, a TSLP inducer. Topical application of MC903 to ear skin increased the expression of not only TSLP but also CCL17, CCL22, the Th2 cytokine IL-4, and the Th17 cytokine IL-17A. Consistently, MC903 induced AD-like skin lesions as shown by increased epidermal thickness; increased infiltration of eosinophils, mast cells, type 2 innate lymphoid cells, Th2 cells, and Th17 cells; and elevated serum levels of total IgE. We also found increased expansion of Th2 cells and Th17 cells in the regional lymph nodes (LNs) of AD mice. Compound 22, a CCR4 inhibitor, ameliorated AD-like skin lesions with reduction of Th2 cells and Th17 cells in the skin lesions and regional LNs. We further confirmed that compound 22 diminished the expansion of Th2 cells and Th17 cells in the coculture of CD11c+ dendritic cells (DCs) and CD4+ T cells derived from the regional LNs of AD mice. Collectively, CCR4 antagonists may exhibit anti-allergic effects by inhibiting both the recruitment and expansion of Th2 cells and Th17 cells in AD.

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

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

CCR4 plays a pivotal role in Th17 cell recruitment and expansion in a mouse model of rheumatoid arthritis.

T helper 17 (Th17) cells express CC chemokine receptor 4 (CCR4) and secrete cytokines such as interleukin-17A (IL-17A) and granulocyte macrophage colony-stimulating factor (GM-CSF), while dendritic cells (DCs) produce CC chemokine ligand 22 (CCL22), a CCR4 ligand, upon stimulation with GM-CSF. Th17 cells are known to play a critical role in the pathogenesis of rheumatoid arthritis (RA). CCL22 has also been shown to be up-regulated in the synovial tissues of RA patients. Here, we investigated the role of CCR4 in collagen-induced arthritis (CIA), a mouse model of RA. DBA/1J mice efficiently developed CIA as shown by erythema, paw swelling, joint rigidity, and joint destruction. Th17 cells were increased in the arthritic joints and regional lymph nodes (LNs) of CIA mice. A fraction of Th17 cells were also shown to produce GM-CSF. On the other hand, we observed no significant increases of Th2 cells or Treg cells, the T cell subsets also known to express CCR4, in these tissues. We further observed clusters of CCR4-expressing memory Th17 cells and CCL22-producing DCs in the regional LNs of CIA mice, supporting the role of the CCR4-CCL22 axis in the expansion of Th17 cells in the regional LNs. Compound 22, a CCR4 inhibitor, ameliorated the disease severity with reduction of Th17 cells in the arthritic joints and regional LNs and Th17-DC clusters in the regional LNs. We further confirmed that CCR4-deficient mice in the C57BL/6J background were highly resistant to CIA induction compared with wild-type mice. Collectively, CCR4 contributes to the pathogenesis of CIA and may thus represent a new therapeutic target for RA.

Anti-allergic effect of ascorbic acid derivative DDH-1 in a mouse model of atopic dermatitis.

Atopic dermatitis (AD) is the most common inflammatory skin disease, which is characterized by excessive Th2 immune responses. In AD patients, the expression of the chemokines CCL17 and CCL22 is increased in skin lesions, leading to the infiltration of Th2 cells. In addition, typical pro-inflammatory cytokines, including TNF-α, IL-1ß and IL-6, have also been shown to be associated with the pathogenesis of AD. Recently, DDH-1, an ascorbic acid derivative, has been synthesized and demonstrated to have a more stabilized structure and better skin penetrability. Furthermore, DDH-1 has been shown to suppress pro-inflammatory cytokine expression in vitro and in vivo. Therefore, using an AD mouse model, we evaluated the effect of DDH-1 to reduce allergic skin inflammation. We found that cutaneous administration of DDH-1 significantly reduced the expression levels of TNF-α, IL-1ß and IL-6 in the skin lesions of AD-like mice. Additionally, DDH-1 administration also significantly reduced the expression levels of CCL17 and CCL22, resulting in decreased skin infiltration of Th2 cells. Consequently, DDH-1 reduced ear and epidermal thickness, the serum IgE levels and the number of infiltrating inflammatory cells and mast cells into the AD-like skin lesions. Combination treatment with DDH-1 and corticosteroid more efficiently improved the skin lesions compared with corticosteroid alone. Collectively, our results suggest that DDH-1 has an anti-allergic effect in an AD mouse model by reducing not only the pro-inflammatory cytokine expression but also the Th2-associated chemokine expression. Thus, DDH-1 may be beneficial for AD treatment and prevention as a monotherapy or in combination with corticosteroids.

Recent Progress in Dendritic Cell-Based Cancer Immunotherapy.

Cancer immunotherapy aims to treat cancer by enhancing cancer-specific host immune responses. Recently, cancer immunotherapy has been attracting much attention because of the successful clinical application of immune checkpoint inhibitors targeting the CTLA-4 and PD-1/PD-L1 pathways. However, although highly effective in some patients, immune checkpoint inhibitors are beneficial only in a limited fraction of patients, possibly because of the lack of enough cancer-specific immune cells, especially CD8+ cytotoxic T-lymphocytes (CTLs), in the host. On the other hand, studies on cancer vaccines, especially DC-based ones, have made significant progress in recent years. In particular, the identification and characterization of cross-presenting DCs have greatly advanced the strategy for the development of effective DC-based vaccines. In this review, we first summarize the surface markers and functional properties of the five major DC subsets. We then describe new approaches to induce antigen-specific CTLs by targeted delivery of antigens to cross-presenting DCs. In this context, the chemokine receptor XCR1 and its ligand XCL1, being selectively expressed by cross-presenting DCs and mainly produced by activated CD8+ T cells, respectively, provide highly promising molecular tools for this purpose. In the near future, CTL-inducing DC-based cancer vaccines may provide a new breakthrough in cancer immunotherapy alone or in combination with immune checkpoint inhibitors.

CCR4 Involvement in the Expansion of T Helper Type 17 Cells in a Mouse Model of Psoriasis.

Psoriasis is a chronic skin disease associated with T helper (Th)17-mediated inflammation. Because CCR4 is a major chemokine receptor expressed on Th17 cells, we investigated the role of CCR4 in a modified imiquimod-induced psoriasis model that showed enhanced skin infiltration of Th17 cells. CCR4-deficient mice had less severe skin disease than wild-type mice. Th17 cells were decreased in the skin lesions and regional lymph nodes of CCR4-deficient mice. In the regional lymph nodes of wild-type mice, CD44+ memory Th17 cells expressing CCR4 were found to be clustered with dendritic cells expressing CCL22, a ligand for CCR4. Such dendritic cell‒Th17 cell clusters were significantly decreased in CCR4-deficient mice. Similar results were obtained using the IL-23‒induced psoriasis model. In vitro, compound 22, a CCR4 antagonist, significantly reduced the expansion of Th17 cells in the coculture of CD11c+ dendritic cells and CD4+ T cells separately prepared from the regional lymph nodes of wild-type mice with psoriasis. In vivo, compound 22 ameliorated the psoriasis-like skin disease in wild-type mice with significant decreases of Th17 cells in the regional lymph nodes and skin lesions. Collectively, CCR4 is likely to play a role in the pathogenesis of psoriasis through the expansion of Th17 cells.

P2X receptor agonist enhances tumor-specific CTL responses through CD70+ DC-mediated Th17 induction.

Extracellular ATP is known to promote Th17 cell differentiation in the intestinal lamina propria by stimulating CD70+CD11clow dendritic cells (DCs) via P2X receptors (P2XRs). Recent studies have also shown that Th17 cells enhance antitumor immunity by directly promoting proliferation of cytotoxic T lymphocytes (CTLs). These finding led us to test a P2XR agonist, αß-methylene ATP (αß-ATP), as a mucosal vaccine adjuvant to promote CTL responses through Th17 induction. We demonstrated that (i) CD70+CD11clow DCs were present in the nasal lamina propria and expressed P2X1R, P2X2R and P2X4R; (ii) CD70+CD11clow DCs isolated from the nasal lamina propria enhanced Th17 cell differentiation of cocultured splenic CD4+ T cells upon stimulation with αß-ATP; (iii) mice intranasally immunized with ovalbumin (OVA) and αß-ATP had increased OVA-specific Th17 cells and CTLs in the nasal lamina propria and regional lymph nodes; (iv) mice intranasally immunized with OVA and αß-ATP also had elevated resistance to E.G7-OVA tumor growth compared with those intranasally immunized with OVA alone; (v) suramin, a broad-range inhibitor of P2 receptors, suppressed the increases of OVA-specific Th17 cells and CTLs in mice intranasally immunized with OVA and αß-ATP; and (vi) suramin also abrogated the enhanced antitumor immunity of mice intranasally immunized with OVA and αß-ATP against E.G7-OVA. Collectively, αß-ATP may be a promising mucosal adjuvant that promotes antigen-specific CTL responses via CD70+CD11clow DC-mediated Th17 induction.

Antitumor effects of novel mAbs against cationic amino acid transporter 1 (CAT1) on human CRC with amplified CAT1 gene.

Copy number alterations detected by comparative genomic hybridization (CGH) can lead to the identification of novel cancer-related genes. We analyzed chromosomal aberrations in a set of 100 human primary colorectal cancers (CRCs) using CGH and found a solute carrier (SLC) 7A1 gene, which encodes cationic amino acid transporter 1 (CAT1) with 14 putative transmembrane domains, in a chromosome region (13q12.3) with a high frequency of gene amplifications. SLC7A1/CAT1 is a transporter responsible for the uptake of cationic amino acids (arginine, lysine, and ornithine) essential for cellular growth. Microarray and PCR analyses have revealed that mRNA transcribed from CAT1 is overexpressed in more than 70% of human CRC samples, and RNA interference-mediated knockdown of CAT1 inhibited the cell growth of CRCs. Rats were immunized with rat hepatoma cells expressing CAT1 tagged with green fluorescent protein (GFP), and rat splenocytes were fused with mouse myeloma cells. Five rat monoclonal antibodies (mAbs) (CA1 ~ CA5) reacting with HEK293 cells expressing CAT1-GFP in a GFP expression-dependent manner were selected from established hybridoma clones. Novel anti-CAT1 mAbs selectively reacted with human CRC tumor tissues compared with adjacent normal tissues according to immuno-histochemical staining and bound strongly to numerous human cancer cell lines by flow cytometry. Anti-CAT1 mAbs exhibited internalization activity, antibody-dependent cellular cytotoxicity, and migration inhibition activity against CRC cell lines. Furthermore, CA2 inhibited the in vivo growth of human HT29 and SW-C4 CRC tumors in nude mice. This study suggested CAT1 to be a promising target for mAb therapy against CRCs.

Transcutaneous immunization with a highly active form of XCL1 as a vaccine adjuvant using a hydrophilic gel patch elicits long-term CD8+ T cell responses.

Memory CD8+ cytotoxic T-lymphocytes (CTLs) play a key role in protective immunity against infection and cancer. However, the induction of memory CTLs with currently available vaccines remains difficult. The chemokine receptor XCR1 is predominantly expressed on CD103+ cross-presenting dendritic cells (DCs). Recently, we have demonstrated that a high activity form of murine lymphotactin/XCL1 (mXCL1-V21C/A59C), a ligand of XCR1, can induce antigen-specific memory CTLs by increasing the accumulation of CD103+ DCs in the vaccination site and the regional lymph nodes. Here, we combined a hydrophilic gel patch as a transcutaneous delivery device and mXCL1-V21C/A59C as an adjuvant to further enhance memory CTL responses. The transcutaneous delivery of ovalbumin (OVA) and mXCL1-V21C/A59C by the hydrophilic gel patch increased CD103+ DCs in the vaccination site and the regional lymph nodes for a prolonged period of time compared with the intradermal injection of OVA and mXCL1-V21C/A59C. Furthermore, the hydrophilic gel patch containing OVA and mXCL1-V21C/A59C strongly induced OVA-specific memory CTLs and efficiently inhibited the growth of OVA-expressing tumors more than the intradermal injection of OVA and mXCL1-V21C/A59C. Collectively, this type of hydrophilic gel patch and a high activity form of XCL1 may provide a useful tool for the induction of memory CTL responses.

Road Dust as a Significant Radiocesium Transporter from Land to River.

This study made it clear that road dust plays an important role for Cs-137 dynamics emitted by the Fukushima Daiichi nuclear power plant accident. It was proved from the Cs-137 and heavy metals determination in road dust, drainage gutter sediment beside pavement, and riverbed sediment around the inflow point of the gutter. Road dust and drainage gutter sediment contained significantly higher concentrations of Cs-137 and Cr, Ni, Cu, Zn, Mo, Cd, Sn, Sb, and Pb than riverbed sediment. These heavy metals are typically enriched in road dust in general and originate in anthropogenic sources. Concentrations of Cs-137 and the heavy metals were higher in riverbed sediments at the inflow point of drainage than in non-inflow points. Drainage gutter sediments exhibited Cs-137 and heavy metal accumulation at the downmost of the gutter, which is the inflow point into the river. Accordingly, distribution of Cs-137 and the heavy metals concentrations were consistent with each other. Moreover, the concentrations of Cs-137 and heavy metals were correlated positively and significantly, with different proportions between sampling sites but similar between sample type and survey date. This indicates that the discharge of Cs-137 and heavy metals is characteristic of the features of the locations, such as Cs-137 and heavy metals concentrations, (micro-) topography, structure of the road and gutter, pavement area, traffic density, and so on. We conclude that road dust is a major medium of Cs-137 transport from land into aquatic ecosystems.

[IgA vasculitis with massive hemorrhage from the jejunum after steroid administration].

A 45-year-old Japanese man presenting with leg purpura, abdominal pain, and arthralgia was diagnosed with IgA vasculitis. His symptoms resolved after the intravenous administration of prednisolone. However, on day 20 of admission, he experienced bloody discharge and hypovolemic shock. The bleeding point was not identified on contrast-enhanced computed tomography scanning. The blood loss was approximately 10800ml and the patient received transfusions of 48 units of concentrated red blood cells, 18 units of fresh frozen plasma, and 30 units of concentrated platelets. Laparotomy and enteroscopy were performed through the incision of the jejunum to detect the bleeding source. Spurting bleeding was observed during the enteroscopy and partial resection of the jejunum was performed. Histopathological examination of the resected specimen revealed large vessels beneath the jejunal ulcer scar, suggesting bleeding from a Dieulafoy's lesion. Leukocytoclastic vasculitis or cytomegalovirus infection was not observed in the resected specimen. Gastrointestinal symptoms in patients with IgA vasculitis usually improve with bowel rest and conservative treatment. Administration of steroids or factor XIII is recommended for patients with severe abdominal pain refractory to conservative management. Rarely, massive bleeding, perforation, intussusception, and/or intestinal obstruction occur in the gastrointestinal tract and these complications affect patients' prognoses. The clinical course in the present patient indicated that severe bleeding from the gastrointestinal tract can occur even after symptom remission in patients with IgA vasculitis. In such cases, prompt treatment, including laparotomy and/or enteroscopy, is essential.

Anti-tumor effects of an antagonistic mAb against the ASCT2 amino acid transporter on KRAS-mutated human colorectal cancer cells.

KRAS mutations are detected in numerous human cancers, but there are few effective drugs for KRAS-mutated cancers. Transporters for amino acids and glucose are highly expressed on cancer cells, possibly to maintain rapid cell growth and metabolism. Alanine-serine-cysteine transporter 2 (ASCT2) is a primary transporter for glutamine in cancer cells. In this study, we developed a novel monoclonal antibody (mAb) recognizing the extracellular domain of human ASCT2, and investigated whether ASCT2 can be a therapeutic target for KRAS-mutated cancers. Rats were immunized with RH7777 rat hepatoma cells expressing human ASCT2 fused to green fluorescent protein (GFP). Splenocytes from the immunized rats were fused with P3X63Ag8.653 mouse myeloma cells, and selected and cloned hybridoma cells secreting Ab3-8 mAb were established. This mAb reacted with RH7777 transfectants expressing ASCT2-GFP proteins in a GFP intensity-dependent manner. Ab3-8 reacted with various human cancer cells, but not with non-cancer breast epithelial cells or ASCT2-knocked out HEK293 and SW1116 cells. In SW1116 and HCT116 human colon cancer cells with KRAS mutations, treatment with Ab3-8 reduced intracellular glutamine transport, phosphorylation of AKT and ERK, and inhibited in vivo tumor growth of these cells in athymic mice. Inhibition of in vivo tumor growth by Ab3-8 was not observed in HT29 colon and HeLa uterus cancer cells with wild-type KRAS. These results suggest that ASCT2 is an excellent therapeutic target for KRAS-mutated cancers.

[Chronic Activation of the Dopaminergic Neuronal Pathway Improves Behavioral Abnormalities in the Prenatal Valproic Acid Exposure Mouse Model of Autism Spectrum Disorder].

Over the last decade there has been an increase in the prevalence of autism spectrum disorder (ASD); however, its pathogenic mechanisms remain unclear. To date, no effective drug has been developed to treat the core symptoms of ASD, especially social interaction deficits. Previous studies have mainly focused on the glutamatergic, GABAergic, and serotonergic signaling pathways; however, a growing number of studies have reported abnormalities in the dopaminergic pathway, such as mutations and functional alterations of dopamine-related molecules, in ASD patients. Furthermore, atypical antipsychotic drugs risperidone and aripiprazole are prescribed for the treatment of non-core symptoms, such as irritability, in patients with ASD. These observations suggest that the dopaminergic pathway is involved in the pathogenesis of ASD. Previously, we have established a mouse model of ASD based on clinical research, which shows that exposure to valproic acid, an antiepileptic drug, during pregnancy causes an increase in the risk of developing ASD in children. This review summarizes our recent studies, which have assessed alterations in the prefrontal dopaminergic pathway. In addition, we discuss the effects of treatment with attention deficit/hyperactivity disorder drugs and atypical antipsychotic drugs, which activate the prefrontal dopaminergic pathway, on ASD-like behavioral abnormalities in the valproic acid exposure mouse model of ASD.

A CCR4 antagonist ameliorates atopic dermatitis-like skin lesions induced by dibutyl phthalate and a hydrogel patch containing ovalbumin.

CCR4 is a chemokine receptor highly expressed by Th2 cells, and regarded as a potential therapeutic target for atopic dermatitis (AD). CCL17 and CCL22 are the CCR4 ligands, and thymic stromal lymphopoietin (TSLP) is shown to promote the expression of CCL17 and CCL22 by dendritic cells. Here, by using dibutyl phthalate (DBP), a TSLP inducer, and a hydrogel patch as a transcutaneous delivery device for ovalbumin, we developed a novel murine AD model and investigated the effect of Compound 22, a CCR4 antagonist. We first found that the mRNA expression of TSLP together with CCL17 and CCL22 was increased in the skins treated with DBP. Furthermore, the topical application of ovalbumin and DBP efficiently and rapidly induced AD-like skin lesions in BALB/c mice, which were characterized by ear swelling accompanied by infiltration of eosinophils, mast cells, and CCR4-expressing Th2 cells in the skin lesions, and elevated total IgE levels in the sera. Using this AD model, we demonstrated that cutaneous administration of Compound 22 inhibited Th2 cell infiltration and ameliorated the AD-like skin lesions. These results suggest that our AD model could be useful for studying new therapeutic strategies. Collectively, CCR4 antagonists may be a promising approach for treating AD.

mS-11, a mimetic of the mSin3-binding helix in NRSF, ameliorates social interaction deficits in a prenatal valproic acid-induced autism mouse model.

Growing evidence suggests pivotal roles for epigenetic mechanisms in both animal models of and individuals with autism spectrum disorders (ASD). Neuron-restrictive silencer factor (NRSF) binds to neuron-restrictive silencing elements in neuronal genes and recruits co-repressors, such as mSin3, to epigenetically inhibit neuronal gene expression. Because dysregulation of NRSF is related to ASD, here we examined the effects of mS-11, a chemically optimized mimetic of the mSin3-binding helix in NRSF, on the behavioral and morphological abnormalities found in a mouse model of valproic acid (VPA)-induced ASD. Chronic treatment with mS-11 improved prenatal VPA-induced deficits in social interaction. Additionally, we found that NRSF mRNA expression was greater in the somatosensory cortex of VPA-exposed mice than of controls. Agreeing with these behavioral findings, mice that were prenatally exposed to VPA showed lower dendritic spine density in the somatosensory cortex, which was reversed by chronic treatment with mS-11. These findings suggest that mS-11 has the potential for improving ASD-related symptoms through inhibition of mSin3-NRSF binding.

Anti-tumor effects of mAb against L-type amino acid transporter 1 (LAT1) bound to human and monkey LAT1 with dual avidity modes.

L-Type amino acid transporter 1 (LAT1) disulfide linked to CD98 heavy chain (hc) is highly expressed in most cancer cells, but weakly expressed in normal cells. In the present study, we developed novel anti-LAT1 mAbs and showed internalization activity, inhibitory effects of amino acid uptake and cell growth and antibody-dependent cellular cytotoxicity, as well as in vivo antitumor effects in athymic mice. Furthermore, we examined the reactivity of mAbs with LAT1 of Macaca fascicularis to evaluate possible side-effects of antihuman LAT1 mAbs in clinical trials. Antihuman LAT1 mAbs reacted with ACHN human and MK.P3 macaca kidney-derived cells, and this reactivity was significantly decreased by siRNAs against LAT1. Macaca LAT1 cDNA was cloned from MK.P3, and only two amino acid differences between human and macaca LAT1 were seen. RH7777 rat hepatoma and HEK293 human embryonic kidney cells expressing macaca LAT1 were established as stable transfectants, and antihuman LAT1 mAbs were equivalently reactive against transfectants expressing human or macaca LAT1. Dual (high and low) avidity modes were detected in transfectants expressing macaca LAT1, MK.P3, ACHN and HCT116 human colon cancer cells, and KA values were increased by anti-CD98hc mAb, suggesting anti-LAT1 mAbs detect an epitope on LAT1-CD98hc complexes on the cell surface. Based on these results, LAT1 may be a promising anticancer target and Macaca fascicularis can be used in preclinical studies with antihuman LAT1 mAbs.

A Highly Active Form of XCL1/Lymphotactin Functions as an Effective Adjuvant to Recruit Cross-Presenting Dendritic Cells for Induction of Effector and Memory CD8+ T Cells.

The chemokine receptor XCR1 is known to be selectively expressed by cross-presenting dendritic cells (DCs), while its ligand XCL1/lymphotactin is mainly produced by activated CD8+ T cells and natural killer cells. Recent studies have shown that XCL1-antigen fusion proteins efficiently induce CD8+ T cell responses by preferentially delivering antigens to XCR1+ DCs. However, XCL1 per se was found to be a poor adjuvant for induction of CD8+ T cell responses. XCL1 is unique because of its lack of one of the two disulfide bonds commonly conserved in all other chemokines and thus has an unstable structure with a relatively weak chemokine activity. In the present study, we generated a variant form of murine XCL1 termed mXCL1-V21C/A59C that contained a second disulfide bond to stabilize its chemokine structure. We confirmed that mXCL1-V21C/A59C had much more potent chemotactic and calcium mobilization activities than the wild type XCL1 (mXCL1-WT). Intradermal injection of mXCL1-V21C/A59C, but not that of mXCL1-WT, significantly increased the accumulation of XCR1+CD103+ DCs in the injection site, and most of the accumulated XCR1+CD103+ DCs were found to take up co-injected ovalbumin (OVA). Furthermore, recruited XCR1+CD103+ DCs efficiently migrated to the draining lymph nodes and stayed for a prolonged period of time. Consequently, mXCL1-V21C/A59C strongly induced OVA-specific CD8+ T cells. The combination of OVA and mXCL1-V21C/A59C well protected mice from E.G7-OVA tumor growth in both prophylactic and therapeutic protocols. Finally, memory CTL responses were efficiently induced in mice immunized with OVA and mXCL1-V21C/A59C. Although intradermal injection of OVA and polyinosinic-polycytidylic acid (poly(I:C)) as an adjuvant also induced CD8+ T cell responses to OVA, poly (I:C) poorly recruited XCR1+CD103+ DCs in the injection site and failed to induce significant memory CTL responses to OVA. Collectively, our findings demonstrate that a highly active form of XCL1 is a promising vaccine adjuvant for cross-presenting DCs to induce antigen-specific effector and memory CD8+ T cells.

Ascorbic acid derivative DDH-1 ameliorates psoriasis-like skin lesions in mice by suppressing inflammatory cytokine expression.

Psoriasis is a chronic inflammatory skin disease in which inflammatory cytokines play a major role in its pathogenesis. Because DDH-1, a novel amphipathic ascorbic acid derivative, has been recently shown to reduce inflammatory cytokine expression in human keratinocytes in vitro, we investigated its effect on imiquimod-induced psoriasis-like skin lesions in C57BL/6 mice. We first found that IL-1ß and TNF-α mRNA expression was significantly decreased in the skin lesions treated with DDH-1. Furthermore, cutaneous administration of DDH-1 ameliorated psoriasis-like skin lesions. These results suggest that DDH-1 may be effective in the prevention and supplemental treatment of psoriasis.