Ionic Liquid-Based Transcutaneous Peptide Antitumor Vaccine: Therapeutic Effect in a Mouse Tumor Model.

Traditional vaccinations need to be injected with needles, and since some people have a strong aversion to needles, a needle-free alternative delivery system is important. In this study, we employed ionic liquids (ILs) for transcutaneous delivery of cancer antigen-derived peptides to obtain anticancer therapeutic effects in a needle-free manner. ILs successfully increased the in vitro skin permeability of a peptide from Wilms tumor 1 (WT1), one of the more promising cancer antigens, plus or minus an adjuvant, resiquimod (R848), a toll-like receptor 7 agonist. In vivo studies demonstrated that concomitant transcutaneous delivery of WT1 peptide and R848 by ILs induced WT1-specific cytotoxic T lymphocyte (CTL) in mice, resulting in tumor growth inhibition in Lewis lung carcinoma-bearing mice. Interestingly, administrating R848 in ILs before WT1 peptides in ILs increased tumor growth inhibition effects compared to co-administration of both. We found that the prior application of R848 increased the infiltration of leukocytes in the skin and that subsequent delivery of WT1 peptides was more likely to induce WT1-specific CTL. Furthermore, sequential immunization with IL-based formulations was applicable to different types of peptides and cancer models without induction of skin irritation. IL-based transcutaneous delivery of cancer antigen-derived peptides and adjuvants, either alone or together, could be a novel approach to needle-free cancer therapeutic vaccines.

Complement activation induced by PEG enhances humoral immune responses against antigens encapsulated in PEG-modified liposomes.

Splenic marginal zone B (MZ-B) cells have attracted attention as alternative antigen-presenting cells. We recently developed an original delivery system, using PEGylated liposomes (PEG-Lip) to deliver antigens to MZ-B cells. In this system, to induce antigen-specific immunity, empty PEG-Lip and antigen-containing PEG-Lip were intravenously (i.v.) injected sequentially at 3 day intervals. Since complement activation by the second dose is required for the delivery of antigen-containing PEG-Lip to splenic MZ-B cells, we investigated the ability of liposomes, modified with various PEG derivatives having different functional terminal groups (methoxy PEG (CH3O-PEG), hydroxy PEG (HO-PEG) or polyglycerol (PG), to activate the complement system and deliver a model antigen, ovalbumin (OVA), to splenic MZ-B cells in vitro and in vivo. Hydroxy PEG-modified liposomes (HO-PEG-Lip) both activated the complement system in vitro, and facilitated the preferential association of HO-PEG-lip with MZ-B cells in vitro. Manipulating HO-PEG density, in particular a density of 2 mol% in total lipids, significantly enhanced the association of HO-PEG-Lip with splenic MZ-B cells in vivo. Consequently, a single i.v. injection of HO-PEG-Lip (2 mol%) containing OVA induced OVA-specific IgG response. Our immunization system with HO-PEG-Lip, achieved efficient antigen delivery to MZ-B cells after a single i.v. injection, improving on our previous immunization system. This new delivery technique may be an improved, simple, antigen delivery system to MZ-B cells that induces meaningful levels of humoral immune response.

Evaluation of the hemodynamic impact of continuous renal replacement therapy in healthy dogs.

We performed continuous renal replacement therapy (CRRT) on clinically healthy dogs to evaluate the effects of CRRT on hemodynamics. Heart rate, arterial blood pressure, and central venous pressure of the dogs (n=6) were recorded during the procedure, which was performed under general anesthesia. Throughout the CRRT, heart rate and arterial blood pressure were stable. Central venous pressure increased after CRRT termination but returned to the basal level within 30 min. In this study, hemodynamic alterations, including hypotension, hypertension, and arrhythmias, were not observed during CRRT. These observations suggest that the CRRT protocol used in the present study can be safely applied to clinical cases with acute renal failure.

Nafamostat mesilate is not appropriate as an anticoagulant during continuous renal replacement therapy in dogs.

We performed continuous renal replacement therapy (CRRT) in clinically healthy dogs (n=7) to evaluate the utility of nafamostat mesilate (NM) as an anticoagulant. In 3 of the 7 dogs, CRRT had to be discontinued before the target duration due to coagulation in the extracorporeal circuit, into which NM was administered constantly at the rate of 2.0-6.0 mg/kg per hour. The rate of administration of NM was greater than the recommended dose of NM in humans. Further, all the dogs suffered vomiting during CRRT with NM infusion. We therefore recommend that NM is not used as an anticoagulant during CRRT in dogs.

Superficial necrolytic dermatitis associated with extrapancreatic glucagonoma in a dog.

An 11-year-old Shih Tzu presented with crusting and erythema, mainly on the abdomen and the root of the tail. Based on histopathological findings, blood examinations and necropsy findings, the condition was diagnosed as superficial necrolytic dermatitis associated with a glucagon-secreting extrapancreatic neuroendocrine tumour. Gross necropsy revealed tumour invasion into the spleen, liver, adrenal glands and mesenteric lymph nodes. Immunohistochemical analysis of the neoplastic cells revealed that the tumour was a glucagonoma, consistent with earlier findings of persistent glucagonaemia and hypoaminoacidaemia.

Placental folate transport during pregnancy.

The aim of this study was to elucidate the mechanism of folate transport in the placenta over the course of pregnancy. We found that folate receptor alpha (FRalpha) and reduced folate carrier (RFC) localized on the apical side of human placental villi. Since folate binding to placental brush-border membrane vesicles (BBMVs) was strongly inhibited by phosphatidylinositol-specific phospholipase C (PI-PLC) treatment, it is possible that FRalpha, a glycosyl phosphatidylinositol linked glycoprotein, is a candidate for folate uptake from maternal blood to the placenta. Moreover, additional inhibitory effects of thiamine pyrophosphate (TPP) and hemin on folate uptake after PI-PLC treatment suggested that not only FRalpha but also RFC and heme carrier protein 1 (HCP1) are involved in the folate transport mechanism in the human placenta. It was also found that accumulation of folate after intravenous injection increased with the progress of gestation in the rat placenta and the fetus. Furthermore, increases in the expression levels of mRNA of rFRalpha, rRFC, and rHCP1 in the rat placenta during pregnancy were observed. These findings suggest that FRalpha, RFC, and HCP1 are important carriers of folate in the placenta during pregnancy. The results of this study suggest that increases in the expression levels of FRalpha, RFC, and HCP1 in the placenta play an important role in the response to increased need for folate for the placenta and fetus during development with the progress of gestation.