Association of Angiopoietin-2 and Ki-67 Expression with Vascular Density and Sunitinib Response in Metastatic Renal Cell Carcinoma

The Angiopoietin-2 (Ang2, Angpt2) growth factor is a context-dependent antagonist/agonist ligand of the endothelial Tie2 receptor tyrosine kinase and known to promote tumour angiogenesis and metastasis. Angiopoietin antagonists have been tested in clinical cancer trials in combination with VEGF-based anti-angiogenic therapy, including sunitinib, which is widely used as a first-line therapy for metastatic renal cell carcinoma (mRCC). However, little is known about Ang2 protein expression in human tumours and the correlation of tumour Ang2 expression with tumour vascularization, tumour cell proliferation and response to anti-angiogenic therapies. Here, we evaluated, using immunohistochemistry, the expression of Ang2, CD31 and the cell proliferation marker Ki-67 in the primary kidney cancer from 136 mRCC patients, who received first-line sunitinib after nephrectomy. Ang2 protein expression was restrained to RCC tumour vessels, and correlated with tumour vascularization and response to sunitinib. High pre-therapeutic Ang2 expression, and more strongly, combined high expression of both Ang2 and CD31, were associated with a high clinical benefit rate (CBR). Low cancer Ki-67 expression, but not Ang2 or CD31 expression, was associated with favourable progression-free (PFS) and overall survival (OS) as compared to patients with high Ki-67 expression (PFS 6.5 vs. 10.6 months, P = 0.009; OS, 15.7 vs. 28.5 months, P = 0.015). In summary, in this study to investigate endothelial Ang2 in mRCC patients treated with first-line sunitinib, high cancer Ang2 expression was associated with the CBR, but not PFS or OS, whereas low Ki-67 expression was significantly associated with long PFS and OS.     

Intermittent cyclophosphamide in refractory rheumatoid arthritis.

Three patients with refractory rheumatoid arthritis were treated with oral cyclophosphamide; in two cases this was supplemented with pulse treatment with methylprednisolone. Long term remission was induced in all three patients and was sustained until follow up at least nine months after the methylprednisolone was stopped. Leucopenia occurred but resolved when cyclophosphamide was reduced from daily to intermittent dosing. Intermittent treatment with cyclophosphamide, possibly in conjunction with pulses of methylprednisolone, may induce remission in patients with rheumatoid arthritis refractory to other forms of treatment.     

Monocyte procoagulant inducing factor: a lymphokine involved in the T cell-instructed monocyte procoagulant response to antigen

A T cell-derived lymphokine couples the recognition of alloantigens by human T inducer cells to monocyte effector procoagulant activity (PCA). This collaborative cellular response results in expression of the functional tissue factor gene product and initiation of the extrinsic coagulation protease cascade as an inflammatory sequelae to the immune response. We now provide initial characterization of this lymphokine, provide evidence that it is a unique lymphokine, and designate it as monocyte procoagulant inducing factor (MPIF). MPIF was produced by alloantigen-stimulated, nylon wool-purified T cells and was not diminished by irradiation. MPIF active supernatants induced PCA in isolated monocytes, and the effect was not modified by the presence of T cells with the responding monocytes, consistent with a direct effect on the monocyte effector cells rather than an indirect effect via an intermediate accessory T cell. Full expression of PCA by monocytes was complete within 4 to 6 hours. MPIF activity in mixed lymphocyte culture (MLC) supernatants (MLC-SN) was stable at pH 2.0 for 24 hr, but was diminished after exposure to pH 10.5 for 30 min. MPIF activity was stable at 56 degrees C but was labile at 63 degrees C or higher. When characterized by chromatography on Sephadex G-100 superfine at either pH 7.2 or 3.6, the activity was recovered in a major discrete peak of about 55,000 daltons, and minor peaks of activity at about 14,000 and greater than 150,000 daltons. There was no correlation between the presence or concentration of INF-gamma, INF-alpha, IL 1, IL 2, GM-CSF, CSF-1, TNF-alpha, TNF-beta (lymphotoxin), or migration inhibitory factor (MIF) with MPIF activity. Each of the previously defined cytokines was analyzed directly for MPIF activity. INF-gamma, INF-alpha, GM-CSF, TNF-alpha, and CSF-1 did not possess MPIF activity over a wide range of concentrations. IL 1 and IL 2 lacked activity at concentrations present in MLC medium positive for MPIF; however, at higher concentrations each demonstrated slight activity. There was a poor correlation between MPIF and MIF activities in MLC-SN, and the content of MIF was insufficient to account for the expressed level of MPIF activity. The lack of identity of these cytokines with MPIF was supported by selected MLC medium that lacked IL 1, IL 2, and MIF and yet contained high MPIF activity. MPIF was additionally distinguished from IL 1, IL 2, and MIF on the basis of separation in Sephadex G-100 superfine.(ABSTRACT TRUNCATED AT 400 WORDS)     

Site-directed mutagenesis of putative active-site residues of Bacillus stearothermophilus α-amylase

Putative catalytic residues of the thermostable Bacillus stearothermophilus -amylase derived by sequence analysis and computer modeling were tested by site-directed mutagenesis. The conservative mutations produced were Asp-234-Glu, Glu-264-Asp, and Asp-331-Asn. The corresponding amino acids have been proposed to act in acid-base catalysis in the Aspergillus oryzae and porcine pancreatic -amylase. Isoelectric focusing and immunodiffusion studies showed that, although inactive, the mutant proteins have conformations similar to the wild type enzyme. The cause of inactivation is presumably a steric clash or alteration of a catalytic amino acid in the case of Asp-234-Glu and a mutation of a catalytic residue in the mutants Glu-264-Asp and Asp-331-Asn.Abbreviations BStA Bacillus stearothermophilus -amylase - PPA porcine pancreatic -amylase - TAA Aspergillus oryzae -amylase     

Associations of HLA-A, -B and -DRB1 Types with Oral Diseases in Swiss Adults

Human leukocyte antigens (HLA) are crucial components of host defense against microbial challenge but the associations of HLA types with oral infectious diseases have not been studied in detail. This prospective cross-sectional study examined associations of HLA-A, -B and -DRB1 types with common oral diseases in a healthy Swiss adult population. 257 subjects (107 m, 150 f, mean age: 43.5 yr; range: 21–58 yr) with known HLA-A, -B and -DRB1 profiles and comprehensive medical records were included. A thorough anamnesis was followed by oral examinations including saliva flow measurements, the DMFT score for cariological status, complete periodontal status with plaque and bleeding indexes as well as assessment of mucosal alterations and temporomandibular dysfunction (TMD). Student’s t-test and Pearson chi-square test were utilized to compare the oral diseases between HLA positive and negative subjects. Bonferroni correction for multiple comparisons was used and P_Bonf<0.05 was considered statistically significant. HLA types -B15 (P_Bonf = 0.002), -B51 (P_Bonf = 0.02) and -DRB1*12 (P_Bonf = 0.02) were associated with less periodontal disease manifestations. HLA-A32 had a positive association with TMD dysfunction (P_Bonf = 0.012). No other statistically significant associations were observed. In conclusion, HLA types may contribute to the development of oral diseases in generally healthy Caucasian adults.     

Chlorpromazine and apigenin reduce adenovirus replication and decrease replication associated toxicity

BACKGROUND: Adenoviruses can cause severe toxicity in immunocompromised individuals. Although clinical trials have confirmed the potency and safety of selectively oncolytic adenoviruses for treatment of advanced cancers, increasingly effective agents could result in more toxicity and therefore it would be useful if replication could be abrogated if necessary. METHODS: We analyzed the effect of chlorpromazine, an inhibitor of clathrin-dependent endocytosis and apigenin, a cell cycle regulator, on adenovirus replication and toxicity. First, we evaluated the in vitro replication of a tumor targeted Rb-p16 pathway selective oncolytic adenovirus (Ad5/3-Delta24) and a wild-type adenovirus in normal cells, fresh liver samples and in ovarian cancer cell lines. Further, we analyzed the in vitro cell killing efficacy of adenoviruses in the presence and absence of the substances. Moreover, the effect on in vivo efficacy, replication and liver toxicity of the adenoviruses was evaluated. RESULTS: We demonstrate in vitro and in vivo reduction of adenovirus replication and associated toxicity with chlorpromazine and apigenin. Effective doses were well within what would be predicted safe in humans. CONCLUSIONS: Chlorpromazine and apigenin might reduce the replication of adenovirus, which could provide a safety switch in case replication-associated side effects are encountered in patients. In addition, these substances could be useful for the treatment of systemic adenoviral infections in immunosuppressed patients.