Epidemiological study using IgM and IgG antibody titers against SARS-CoV-2 in The University of Tokyo, Japan (UT-CATS).

INTRODUCTION: The worldwide pandemic of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has continued to date. Given that some of the patients with coronavirus disease 2019 (COVID-19) are asymptomatic, antibody tests are useful to determine whether there is a previous infection with SARS-CoV-2. In this study, we measured IgM and IgG antibody titers against SARS-CoV-2 in the serum of asymptomatic healthy subjects in The University of Tokyo, Japan. METHODS: From June 2020, we recruited participants, who were students, staff, and faculty members of The University of Tokyo in the project named The University of Tokyo COVID-19 Antibody Titer Survey (UT-CATS). Following blood sample collection, participants were required to answer an online questionnaire about their social and health information. We measured IgG and IgM titers against SARS-CoV-2 using iFlash-SARS-CoV-2 IgM and IgG detection kit which applies a chemiluminescent immunoassay (CLIA) for the qualitative detection. RESULTS: There were 6609 volunteers in this study. After setting the cutoff value at 10 AU/mL, 32 (0.48%) were positive for IgG and 16 (0.24%) for IgM. Of six participants with a history of COVID-19, five were positive for IgG, whereas all were negative for IgM. The median titer of IgG was 0.40 AU/mL and 0.39 AU/mL for IgM. Both IgG and IgM titers were affected by gender, age, smoking status, and comorbidities. CONCLUSIONS: Positive rates of IgG and IgM titers were relatively low in our university. Serum levels of these antibodies were affected by several factors, which might affect the clinical course of COVID-19.

Establishment of a stable sampling method to assay mercaptoalbumin/non-mercaptoalbumin and reference ranges.

Oxidative stress is reportedly associated with many diseases such as cancer, arteriosclerosis, diabetes and aging, but no practical biomarkers are currently available in actual clinical practice. Human mercaptoalbumin (HMA) and human non-mercaptoalbumin (HNA) are expected to become markers of oxidative stress, but the stability of HMA/HNA has been problematic. We investigated the conditions for stabilizing HMA/HNA and found that HMA/HNA was stable at room temperature for 25 h if whole blood samples were mixed with a citrate buffer so that the citric acid concentration after mixing was 70 mM or higher and the pH of the added buffer was less than pH 6.0. Whole blood samples were then collected under the above conditions, and the reference range for HNA was set at 21.8% ±â€¯7.4% (HMA, 78.2% ±â€¯7.4%) based on samples from 65 volunteers (28 males and 37 females; average age, 55.0 ±â€¯13.8 years). The clinical usefulness of HMA/HNA as an oxidative stress marker should be clarified for specific pathological conditions using the previously reported, highly accurate measurement method under the conditions required for HMA/HNA stability.

Autotaxin and soluble IL-2 receptor concentrations in cerebrospinal fluids are useful for the diagnosis of central nervous system invasion caused by haematological malignancies.

BACKGROUND: Invasion of the central nervous system by haematological malignancies is diagnosed by cytological analyses of cerebrospinal fluid or diagnostic imaging, while quantitative biomarkers for central nervous system invasion are not available and needed to be developed. METHODS: In this study, we measured the concentrations of autotaxin and soluble IL-2 receptor in cerebrospinal fluid and evaluated their usefulness as biomarkers for central nervous system invasion. RESULTS: We observed that both the autotaxin and soluble IL-2 receptor concentrations in cerebrospinal fluid were higher in subjects with central nervous system invasion than in those without, and the cerebrospinal fluid concentrations were independent from the serum concentrations of these biomarkers. ROC analyses revealed that the soluble IL-2 receptor concentration in cerebrospinal fluid was a strong discriminator of central nervous system invasion in subjects with haematological malignancies, while the autotaxin concentration in cerebrospinal fluid also had a strong ability to discriminate central nervous system invasion when the subjects were limited to those with lymphoma. The combined measurement of autotaxin and soluble IL-2 receptor in cerebrospinal fluid improved the sensitivity without notably reducing the specificity for central nervous system invasion in subjects with lymphoma when central nervous system invasion was diagnosed in cases where either value was beyond the respective cut-off value. CONCLUSION: These results suggest the possible usefulness of soluble IL-2 receptor and autotaxin concentrations in cerebrospinal fluid for the diagnosis of central nervous system invasion.

[Evaluation of Immature Platelet Fraction Measurements Using an Automated Hematology Analyzer, Sysmex XN].

The immature platelet fraction (IPF%) is a parameter for the automatic quantification of reticulated plate- lets and is considered to reflect platelet production. We evaluated IPF% measurements using an automated analyzer, the Sysmex XN. We measured the platelet counts and the IPF% values in 35 healthy subjects and 275 patients with various diseases using both the XN analyzer and a conventional analyzer, the Sysmex XE. Significant correlations in the platelet count and the IPF% were observed between the XN results and the XE results. In the same samples, significant inverse correlations were observed between the platelet count and the IPF% in both the XN and XE results. The coefficient of variation values for the platelet count and the IPF% measurements obtained using the XN analyzer were lower than those obtained using the XE ana- lyzer. In patients who had undergone hematopoietic stem cell transplantation, the IPF% measurements obtained using the XN analyzer increased several days before platelet recovery. IPF% measurements performed using the XN analyzer are adequate for clinical use. This parameter may be a useful marker for the prediction of platelet recovery. [Original].

Isoprenoid geranylgeranylacetone inhibits human colon cancer cells through induction of apoptosis and cell cycle arrest.

Geranylgeranylacetone (GGA), an isoprenoid compound, is a widely used antiulcer drug developed in Japan. GGA is structurally similar to plaunotol and geranylgeraniol, another isoprenoid reported to exert strong anticancer effects. In an earlier study, GGA was shown to inhibit ovarian cancer invasion by attenuating not only Rho activation, but also Ras-MAPK activation. In this study, we aimed to test whether GGA could have a therapeutic effect on colon cancer cells. As a result, we found that GGA induced a dose-dependent decrease in the proliferative activity through induction of cell apoptosis and cell cycle arrest in the G1 phase. The induction of apoptosis was mediated by the activation of both caspase-8 and caspase-9 pathways. The induction of G1 arrest was mediated by the increase of p21 and p27, and also the decrease of phosphorylated retinoblastoma protein levels. This study showed the potential anticancer activity of GGA. As this drug is already available in Japan for clinical use as an antiulcer/antigastritis agent, clinical trials will be designed to confirm its potential usefulness for cancer patients.

Id1/Id3 knockdown inhibits metastatic potential of pancreatic cancer.

BACKGROUND: The Id (inhibitor of DNA binding/differentiation) proteins belong to the helix-loop-helix transcriptional regulatory factors, and play important roles in tumor development. Previously, we and others have shown that targeting Id in tumor cells could have important clinical implications. In the present study, we aimed to evaluate the effects of Id inhibition in human pancreatic cancer cells. MATERIALS AND METHODS: Id1 and Id3 were stably double-knockdown in human pancreatic cancer cell line MIA-Paca2 by means of RNA interference. Expression of Id and integrins were analyzed by flow-cytometry. Cell proliferation was evaluated by MTS assay. Migration was measured by wound closure assay. Adhesion assay was performed to evaluate binding capacity for different extracellular matrix proteins. Finally, in vivo properties of tumor cells were observed in a mouse model of peritoneal metastasis. RESULTS: Id1/Id3 double-knockdown resulted in decreased ability of pancreatic cancer cells to proliferate and migrate. In addition, Id1/Id3 double-knockdown caused decreased expression of integrins alpha3, alpha6, and beta1, and consequently reduced adhesion of tumor cells to laminin. Finally, peritoneal metastases of Id1/Id3 double-knockdown tumor cells were significantly reduced. CONCLUSIONS: We concluded that the Id proteins play a pivotal role in the development of peritoneal metastasis of pancreatic cancer, and consequently, their targeting would be a novel strategy for the prevention and treatment of pancreatic cancer.

Plaunotol and geranylgeraniol induce caspase-mediated apoptosis in colon cancer.

BACKGROUND: Plaunotol, a kind of isoprenoid extracted from a Thai medical plant, plau-noi, is structurally similar to geranylgeraniol (GGOH), another isoprenoid reported to exert strong anticancer effects. Recently, we have reported on its inhibitory effects on tumor angiogenesis and direct effects on gastric cancer cells. Here, we aimed to test whether plaunotol could have some therapeutic effect on colon cancer. MATERIALS AND METHODS: Human colon cancer cell line DLD1 was used. Tumor cells were cultured in the presence of plaunotol or GGOH, and their proliferation was measured by MTS assay. Apoptosis was evaluated by Annexin V and propidium iodide double-staining or terminal-deoxynucleotidyl assay. The activation of caspase-3, -8, and -9 was analyzed by flow cytometry and Western blot analysis for PRRP cleavage. RESULTS: Plaunotol and GGOH strongly inhibited the proliferative activity of DLD1, dependent on induction of apoptosis. The induction of apoptosis by either plaunotol or GGOH was dependent on the activation of both caspase-8 and caspase-9 pathways. CONCLUSIONS: Plaunotol would be a potential anticancer agent against colon cancer, and since it is already available in Japan and Thailand for clinical use as an anti-ulcer/antigastritis agent, clinical trials will be designed to confirm the present findings.

Pilot study of anti-angiogenic vaccine using fixed whole endothelium in patients with progressive malignancy after failure of conventional therapy.

Vaccines targeting tumour angiogenesis were recently shown to inhibit tumour growth in animal models. However, there is still a lack of information about the clinical utility of anti-angiogenic vaccination. Therefore, here, we aimed to test the clinical effects of a vaccine using glutaraldehyde-fixed human umbilical vein endothelial cells (HUVECs). Six patients with recurrent malignant brain tumours and three patients with metastatic colorectal cancer received intradermal injections of 5x10(7) HUVECs/dose (in total 230 vaccinations). ELISA and flow cytometry revealed immunoglobulin response against HUVECs' membrane antigens. ELISPOT and chromium-release cytotoxicity assay revealed a specific cellular immune response against HUVECs, which were lysed in an effectors:targets ratio-dependent manner. Gadolinium-contrasted MRI showed partial or complete tumour responses in three malignant brain tumour patients. Except for a DTH-like skin reaction at the injection site, no adverse effect of vaccination could be observed. Our results suggest that the endothelial vaccine can overcome peripheral tolerance of self-angiogenic antigens in clinical settings, and therefore should be useful for adjuvant immunotherapy of cancer.