Adjuvant-mediated tumor regression and tumor-specific cytotoxic response are impaired in MyD88-deficient mice

The Mycobacterium bovis bacillus Calmette-Guérin cell-wall skeleton (BCG-CWS) activates Toll-like receptor (TLR) 2 and TLR4, but unlike the typical TLR4 agonist bacterial lipopolysaccharide barely induces type 1 IFN. BCG-CWS has been used for adjuvant immunotherapy for patients with cancer. We investigated the adjuvant potential of BCG-CWS for induction of CTLs subsequent to TLR-mediated dendritic cell (DC) maturation, using a syngeneic mouse tumor model (B16 melanoma in C57BL/6). We evaluated the retardation of tumor growth and cytotoxic response in wild-type and MyD88-/- mice immunized with tumor debris and/or BCG-CWS. Delays in tumor growth and cytotoxic response were induced by immunization with a mixture of BCG-CWS emulsion and the tumor. BCG-CWS was capable of activating DCs ex vivo by the criteria of CD80/CD86 up-regulation and cytokine (interleukin-12, tumor necrosis factor-alpha) induction. Efficient tumor suppression and ex vivo cytokine induction did not occur in MyD88-deficient mice and cells, suggesting that the MyD88 adapter is crucial for induction of tumor cytotoxicity. Because TLR4 is involved in both MyD88-dependent and -independent pathways and the latter affects DC maturation, our findings indicate that both pathways cooperate to induce CTL-based tumor immunity.     

Characterization of OM-B Monoclonal Antibody-defined Antigen Associated with Mucinous Type Human Ovarian Tumor

Partial characterization of the OM-B antigen associated with mucinous-type ovarian tumors was conducted. This antigen was defined by OM-B monoclonal antibody, which was raised against a mucinous-type ovarian tumor, and was present in all the mucinous-type tumors tested, but only a fraction of serous-type tumors. The OM-B crude antigen preparation fractionated from cystic fluids had a density of 1.40–1.43 g/ml, with a high neutral sugar content. Molecular mass ( M r) estimated by gel filtration was more than 2,000,000. Trypsinization of the antigen preparation under appropriate conditions resulted in two major bands and one minor band with molecular sizes of less than M r 250,000, as detected by immunoblotting. Immunoaffinity chromatography was then conducted and the amino acid composition of the purified product was determined; the high contents of serine, threonine and proline are characteristic of a mucin. Binding inhibition enzyme-linked immunosorbent assay was developed to measure OM-B antigen activity in cystic fluids and sera from patients with mucinous-type tumors. The antigen was easily detected in most cystic fluids, but not in sera, suggesting that improvement in the sensitivity of this assay is necessary before its utilization for serum diagnosis will be feasible.     

Distribution of adenosine deaminase binding protein in normal and malignant tissues of the gastrointestinal tract studied by monoclonal antibodies

A mouse monoclonal antibody V-715 was raised against fresh colon cancer tissues. Biochemical analysis elucidated that the antigen defined was adenosine deaminase binding protein (ADBP). In colon cancer cell lines, V-715 was positive in 8 out of 16 differentiated cancers and in 2 out of 8 poorly differentiated cancers. In frozen sections, ADBP was expressed in 17 out of 33 differentiated colon cancers, but none of 4 poorly differentiated colon cancers. In normal colon, the expression was observed in epithelium. In gastric cancers, ADBP was expressed in 10 out of 15 differentiated cancers, but weakly or only heterogenously expressed in 2 out of 8 poorly differentiated cancers. In normal gastric mucosa, ADBP was mainly detected in the foveolar epithelium, but was weakly or not expressed in the deep gastric glands. Carcinoid tumors and malignant lymphoma of the stomach did not express ADBP. These results suggest that ADBP may act as a marker of enterocytic differentiation in normal and neoplastic gastrointestinal cells, and might be exploitable in clinical and pathological diagnosis of gastrointestinal cancers. © 1993 Wiley-Liss, Inc.     

IFN-β Gene Therapy Induces Systemic Antitumor Immunity Against Malignant Glioma

We previously demonstrated that intratumoral administration of liposomes containing the murine interferon beta (IFN-) gene [lip(pSV2muIFN-)] resulted in stronger growth-inhibitory effect on GL261 (H-2^b) mouse glioma inoculated in brains of syngeneic C57BL/6 mice than conventional exogenous IFN- administration, and histologic evaluation revealed the massive infiltration of T lymphocytes (CD8 > CD4) within the residual tumor. The present study was aimed at determining whether such tumor-infiltrating lymphocytes (TIL) have any tumor-specific cytotoxic effects. Intratumoral administration of lip(pSV2muIFN-) resulted in prolonged survival time and a 50% tumor-free incidence in the mice treated. The surviving animals were subsequently re-challenged with either subcutaneous or intracranial injection of GL261 cells, and no tumors were found to develop over a 50-day period. In vivo depletion of CD8, but not CD4 cells decreased the efficacy of lip(pSV2muIFN-). Specific cytotoxic T lymphocytes (CTL) against GL261 cells were generated from both TIL and spleen cells of the mice treated. The results of flow cytometric analysis and antibody blocking test revealed that the bulk CTL lines thus prepared were T cell receptor (TCR) , CD8 T lymphocytes with H-2^b restriction.These findings suggest that, in addition to direct growth-inhibitory effects by the IFN- gene on the tumor cells, activation of systemic cellular immunity may participate in antitumor effects in vivo, despite the fact that central nervous system is generally regarded as an immunologically privileged site.     

Pulmonary hemodynamics following reconstructive mitral valve surgery

The hemodynamic response in the immediate postoperative period after reconstructive mitral valve surgery was evaluated by means of cardiac output, pulmonary artery and left atrial pressures, pulmonary blood volume and pulmonary vascular resistance using indwelling catheters in the pulmonary artery and left atrium. Elevated pulmonary artery pressure returned to near-normal levels immediately after the correction of mitral stenosis by closed commissurotomy and mitral insufficiency by annuloplasty or valvuloplasty. Pulmonary artery and left atrial pressures returned to near-normal levels within 24 hours after surgery. In contrast, normalization of pulmonary artery and left atrial pressures were not conspicuous in cases of open commissurotomy. The difference could not be explained by the blood volume deficits, ischemic cardiac arrest during surgery, or residual valvular lesions. The most likely explanation appears to be due to further impairment of chronically diseased left ventricle by open heart surgery and increased water load after hemodilution perfusion which leads to interstitial pulmonary edema. Comparison between the pulmonary artery, left atrial pressures and cardiac output suggested that keeping pulmonary artery mean pressure below 35 mmHg. and left atrial mean pressure below 15 mmHg. are important to maintain optimum cardiac output without causing interstitial pulmonary edema in the immediate postoperative period after mitral valve surgery.     

Identification of an HLA-A24-restricted cytotoxic T lymphocyte epitope from human papillomavirus type-16 E6: the combined effects of bortezomib and interferon-gamma on the presentation of a cryptic epitope

About 50% of cervical cancers are associated with human papillomavirus type 16 (HPV-16), and since the HPV-16 E6 and E7 oncoproteins are constitutively expressed in the tumor cells, they are attractive targets for cytotoxic T lymphocyte (CTL)-mediated immunotherapy. Nevertheless, only a limited number of HPV-16 E6 epitopes have been identified to date. Using reverse immunological methods, we have generated a CTL clone against the HPV-16 E6(49-57) epitope restricted by HLA-A*2402, which is the most common allele in Japan and relatively frequent worldwide, capable of lysing 293T cells transduced with HLA-A*2402 and HPV-16 E6. Although it was unable to recognize the SiHa cervical cancer cell line positive for HPV-16 and HLA-A*2402, the cells became susceptible to lysis when transduced with E6-E7 genes, which was unexpectedly offset by pretreatment with interferon (IFN)-gamma alone. Interestingly, however, combined pretreatment with a proteasome inhibitor, bortezomib and IFN-gamma fully restored CTL-mediated lysis of the original SiHa cells. Furthermore, such intervention of 2 of 4 other cervical cancer cell lines expressing HPV-16 E6 and HLA-A*2402 was found to induce IFN-gamma production by specific CTLs. Tetramer analysis further revealed that induction of E6(49-57)-specific T cells was possible in 5 of 7 patients with HPV-16-positive high grade cervical intraepithelial neoplasia or cervical cancer by in vitro stimulation with E6(49-57) peptide. Thus, these findings together indicate that E6(49-57) is a candidate epitope for immunotherapy and immunological monitoring of such patients.     

Molecular Cloning of 19p13 Breakpoint Region in Infantile Leukemia with t(11;19)(q23;p13) Translocation

We studied the breakpoint regions involved in t(11;19)(q23;p13) translocation associated with infantile leukemias. Southern blot analysis with the partial cDNA clone for the MLL gene at 11q23 which we had isolated previously detected gene rearrangements in all three cell lines and three leukemia samples from the patients with t(11;19) translocation, indicating that these breakpoints were clustered within the 8.5 kb Bam HI germline fragment detected by the probe. To study the breakpoint region, a genomic library of one of the cell lines, KOCL-33, was made. We have isolated the der(19) allele containing the breakpoint as well as the germline alleles at 19p13 and 11q23. Using the genomic probes on chromosome 19 near the breakpoint, Southern blot analysis was performed. The breakpoints at 19p13 of the two other cell lines and the three leukemia samples were not located within 36 kilobases of the KOCL-33 breakpoint, although pulsed-field gel electrophoresis showed that the breakpoints of all three cell lines were on the same Nru I fragment of 230 kilobases. These results showed that the breakpoints at 19p13 were not clustered like those at 11q23 in t(11;19) translocation.     

Formalin-treated UV-inactivated SARS coronavirus vaccine retains its immunogenicity and promotes Th2-type immune responses

The demand for rapid and simple development of a vaccine against a newly emerging infectious disease is increasing worldwide. We previously revealed that UV-inactivated severe acute respiratory syndrome (SARS)-associated coronavirus (SARS-CoV) virions (UV-V) elicited high levels of humoral immunity and a weak Th0 response in mice immunized subcutaneously. To ensure the safety of such a whole inactivated SARS-CoV vaccine, we additionally treated the UV-V vaccine with formalin, resulting in the UV-F-V vaccine. Analysis of the immunogenicity of the UV-F-V+alum vaccine in mice revealed that it generated comparable neutralizing serum anti-SARS-CoV IgG antibody levels as the UV-V+alum vaccine. Moreover, both vaccines induced similar frequencies of anti-SARS-CoV IgG antibody-producing cells in bone marrow. Interestingly, the UV-F-V vaccine induced fewer IgG(2a) subtype antibodies and higher interleukin-4 production in vaccinated mice than did UV-V. Thus, UV-F-V imposes a Th2-type bias on the immune response, unlike UV-V. We propose here that doubly-inactivated SARS-CoV virions by UV and formalin constitute a safe vaccine that may effectively induce neutralizing antibodies in humans.     

Responsiveness of the Japanese Orthopaedic Association Back Pain Evaluation Questionnaire in lumbar surgery and its threshold for indicating clinically important differences

Background Context

Introduced in 2007, the Japanese Orthopaedic Association Back Pain Evaluation Questionnaire (JOABPEQ) has been widely used, but its psychometric properties have not been well studied.