Diabody-based recombinant formats of humanized IgG-like bispecific antibody with effective retargeting of lymphocytes to tumor cells

Recently, recombinant antibodies have been dissected into antigen-binding regions and rebuilt into multivalent high-avidity formats. These new structural designs are expected to improve in vivo pharmacokinetics and efficacy in clinical use. Here, we designed effective recombinant bispecific antibody (BsAb) formats based on hEx3, a humanized bispecific diabody with epidermal growth factor receptor and CD3 retargeting. The bispecific and bivalent IgG-like antibodies engineered from hEx3 (or its single-chain form, hEx3-scDb) and the human Fc region showed stronger binding to each target cell than did monovalent diabody formats, and their affinity was identical to that of the corresponding parent IgG. The bivalent effect of the constructed IgG-like BsAbs resulted in cell cytotoxicity 10 times that of monovalent diabodies, and further, the fusion of Fc portion contributed intense cytotoxicity in peripheral blood mononuclear cells by the induction of the antibody-dependent cellular cytotoxicity. The growth-inhibition effects of IgG-like BsAbs were superior to those of the approved therapeutic antibody cetuximab, which recognizes the same epidermal growth factor receptor antigen, even when peripheral blood mononuclear cells were used as effector cells. We thus demonstrated a critical improvement in the effect of hEx3 by the bottom-up construction of IgG-like BsAbs; in adoptive immunotherapy, monotherapy without supplemental molecules may be able to induce antibody-dependent cellular cytotoxicity.     

Highly sensitive upregulation of apolipoprotein A-IV by peroxisome proliferator-activated receptor alpha (PPARalpha) agonist in human hepatoma cells

Peroxisome proliferator-activated receptor alpha (PPARalpha) is a key regulator in hepatic lipid metabolism and a potential therapeutic target for dyslipidemia. However, in humans hepatic PPARalpha-regulated genes remain unclear. To investigate the effect of PPARalpha agonism on mRNA expressions of lipid metabolism-related genes in human livers, a potent PPARalpha agonist, KRP-101 (KRP), was used to treat the human hepatoma cell line, HepaRG cells. KRP did not affect AOX or L-PBE, which are involved in peroxisomal beta-oxidation. KRP increased L-FABP, CPT1A, VLCAD, and PDK4, which are involved in lipid transport or oxidation. However, the EC(50) values (114-2500 nM) were >10-fold weaker than the EC(50) value (10.9 nM) for human PPARalpha in a transactivation assay. To search for more sensitive genes, we determined the mRNA levels of apolipoproteins, apoA-I, apoA-II, apoA-IV, apoA-V, and apoC-III. KRP had no or little effect on apoA-I, apoC-III, and apoA-II. Interestingly, KRP increased apoA-IV (EC(50), 0.99 nM) and apoA-V (EC(50), 0.29 nM) with high sensitivity. We identified apoA-IV as a PPARalpha-upregulated gene in a study using PPARalpha siRNA. Moreover, when administered orally to dogs, KRP decreased the serum triglyceride level and increased the serum apoA-IV level in a dose-dependent manner. These findings suggest that apoA-IV, newly identified as a highly sensitive PPARalpha-regulated gene in human livers, may be one of the mechanisms underlying PPARalpha agonist-induced triglyceride decrease and HDL elevation.     

Associations Between hOGG1 Ser326Cys Polymorphism and Increased Body Mass Index and Fasting Glucose Level in the Japanese General Population

Background

Evidence suggests that Ser326Cys, a genetic polymorphism of human 8-oxoguanine glycosylase 1 (hOGG1), is associated with insulin resistance and type 2 diabetes; however, the underlying mechanism is unclear. Recently, an animal study showed a significant association between the hOGG1 genotype and obesity, although evidence for such an association in humans is limited. The purpose of this study was to examine the association between the hOGG1 genotype and body mass index (BMI) and fasting blood glucose (FBG) levels.

Methods

Cross-sectional analysis was conducted using the baseline survey data from a Japan Multi-Institutional Collaborative Cohort Study, which included 1793 participants aged 40–69 years. The hOGG1 polymorphism was detected using a multiplex polymerase chain reaction-based invader assay. Multiple linear regression, analysis of covariance, and logistic regression were used to control for confounding variables.

Results

The Cys allele was significantly associated with increased BMI, FBG level, and total cholesterol (TC) level, even after adjustment for gender, age, energy intake, alcohol, smoking, physical activity, and family history of diabetes. An association with BMI was still observed after further adjustment for FBG and TC, but not for the study area (Amami or the mainland). The Cys/Cys genotype was significantly more prevalent in the participants with higher BMI (>27.5 kg/m²). However, the impact of genotype decreased and significance disappeared after adjusting for the study area.

Conclusions

The present results suggest that the study area being inside Japan confounds the association between hOGG1 genotype and obesity.Key words: human 8-oxoguanine glycosylase 1 (hOGG1), obesity, body mass index (BMI), fasting blood glucose (FBG), polymorphism, study area     

Epidrum<Superscript>®</Superscript>: a new device to identify the epidural space with an epidural Tuohy needle

Epidrum^® is an optimal pressure, loss of resistance device for identifying the epidural space. We investigated the usefulness of Epidrum versus the loss of resistance or hanging drop techniques while performing epidural anesthesia. Eighty adult patients who were scheduled for elective surgery under lumbar epidural anesthesia were randomized into two groups. The first group (Epidrum group) consisted of 40 adult patients who were scheduled for epidural anesthesia using Epidrum. The second group (control group) consisted of 40 adult patients who were scheduled for epidural anesthesia using the loss of resistance or hanging drop technique. We recorded the time required to identify the epidural space and outcomes of epidural catheterization. The attending anesthesiologists were also questioned regarding the ease of control of the Tuohy needle and of epidural space identification with each method. The time required to perform epidural anesthesia was significantly shorter in the Epidrum group than in the control group [28 s (10–76) vs. 90 s (34–185); median (interquartile range)] (p < 0.05). Tuohy needle control was significantly easier in the Epidrum group than in the control group (p < 0.05). Epidrum is useful for performing epidural anesthesia quickly while obtaining good Tuohy needle control.     

Tumoral calcinosis in bilateral facet joints of the lumbar spine in scleroderma. Case report

Tumoral calcinosis commonly occurs in the articular soft tissues of the extremities but rarely in the spine. The authors performed surgery to treat lumbar tumoral calcinosis in a patient with scleroderma, in whom symptoms of neurological dysfunction had manifested. This 49-year-old woman presented with low-back pain and gait disturbance. Seven years before presentation, scleroderma had been diagnosed, and the patient had received medical treatment ever since. Imaging revealed tumoral calcinosis centered at the bilateral facet joints between L-3 and L-4, marked stenosis of the spinal canal, L-3 spondylolisthesis, and intervertebral instability. Surgery was performed to excise the lesion en bloc. After neural decompression, posterolateral fusion and pedicle screw fixation were undertaken. Symptoms improved after surgery. In this case, the underlying scleroderma that predisposes to calcinosis and facet joint degeneration due to lumbar spondylolisthesis were probably factors leading to the development of tumoral calcinosis in the lumbar spine.     

Hepatic resection through an anterior approach employing a modified liver hanging maneuver in patients with a massive liver tumor severely oppressing the inferior vena cava

For a large hepatic neoplasm existing in the right hepatic lobe, hepatic resection using an anterior approach is required. We have reported an operative procedure for hepatic transection using absorbable polyglycolic acid tape. In patients with suspected tumor invasion of the inferior vena cava, on the other hand, considering the range of the residual tumor while sparing the inferior vena cava as much as possible, combined resection and reconstruction of the inferior vena cava is conducted only if operative curativity is expected. We conducted hepatic transection while maintaining the blood flow of the residual liver by applying the liver hanging maneuver method of Belghiti et al. and polyglycolic acid tape in patients with giant liver tumors of the right hepatic lobe compressing the hepatic inferior vena cava. Strong angled dissecting forceps were inserted into the ventral side of the inferior vena cava from the caudal side, and the tip was induced between hepatic veins. Two strips of polyglycolic acid tape were pinched with forceps and strongly ligated on the right and left sides of the cutoff line. Subsequently, hepatic transection was conducted using electrocautery spray coagulation and CUSA without blocking the inflow blood of the residual liver, and the right hepatic lobe was extirpated. This procedure has already been performed in 5 patients suspected of inferior vena cava invasion, and the inferior vena cava was able to be preserved in all the patients.     

Antibody titer to gp210-C terminal peptide as a clinical parameter for monitoring primary biliary cirrhosis

BACKGROUND/AIMS: The presence of antibodies to the 210-kDa glycoprotein of the nuclear pore complex (gp210) is highly indicative of primary biliary cirrhosis (PBC). However, the significance of anti-gp210 antibody titers for monitoring PBC remains unresolved. METHODS: We used an ELISA with a gp210 C-terminal peptide as an antigen to assess serum antibody titers in 71 patients with PBC. RESULTS: Patients were classified into three groups: Group A in whom anti-gp210 titers were sustained at a high level, Group B in whom anti-gp210 status changed from positive to negative under ursodeoxycholic acid (UDCA) therapy, Group C in whom anti-gp210 antibodies were negative at the time of diagnosis. The rate of progression to end-stage hepatic failure was significantly higher in group A (60%) as compared to groups B (0%) and C (4.2%). The sustained antibody response to gp210 was closely associated with the severity of interface hepatitis. The significance of anti-gp210 antibody was confirmed by National Hospital Organization Study Group for Liver Disease in Japan. CONCLUSIONS: The serial quantitation of serum anti-gp210-C-terminal peptide antibodies is useful for monitoring the effect of UDCA and for the early identification of patients at high risk for end-stage hepatic failure.     

Multiple bone fractures found in a young sarcoidosis patient with long stable disease

A 22-year-old Japanese man was found to have bilateral hilar lymphadenopathy (BHL), and was diagnosed with sarcoidosis in 1995. He was followed without treatment until 2002, when a bone fracture due to osseous sarcoidosis was found in his left thumb. Despite systemic treatment with corticosteroid and methotrexate, a new bone lesion developed in his right foot and his right middle finger was fractured. The patient also suffered multiple organ involvements including brain and muscle lesions. This is the first report of a sarcoidosis patient who presented with BHL, and developed bone fractures after a long stable period of more than 5 years.