Searching one or two databases was insufficient for meta-analysis of observational studies

OBJECTIVE: To address methodologic issues in searching for observational studies by presenting database search methods and results. STUDY DESIGN AND SETTING: Results of two literature searches for publications reporting on observational studies of alcohol consumption and the risk of breast cancer and large bowel cancer were compared, to evaluate the sensitivity of various bibliographic databases and search strategies, including hand-searching reviews and meta-analyses. RESULTS: The target sensitivity of 90% of publications in the breast cancer search was achieved by starting with Medline, then adding Biosis, Embase, and SCI EXPANDED-SSCI, which provided a total of 72 (91%) of the 79 relevant publications. To reach a similar 89% sensitivity for large bowel cancer, at least Biosis, Dissertation Abstracts Online, Embase, ETOH, and Medline had to be searched, with the addition of hand search of reviews and meta-analyses. CONCLUSION: Limiting a search to one or two databases when conducting meta-analyses of observational studies will not provide a thorough summary of the existing literature. The findings support recommendations to implement a comprehensive search of electronic databases and the reference lists of recent review articles and meta-analyses.     

Increased plasma vascular endothelial growth factor (VEGF) as a surrogate marker for optimal therapeutic dosing of VEGF receptor-2 monoclonal antibodies

A major obstacle compromising the successful application of many of the new targeted anticancer drugs, including angiogenesis inhibitors, is the empiricism associated with determining an effective biological/therapeutic dose because many of these drugs express optimum therapeutic activity below the maximum tolerated dose, if such a dose can be defined. Hence, surrogate markers are needed to help determine optimal dosing. Here we describe such a molecular marker, increased plasma levels of vascular endothelial growth factor (VEGF), in normal or tumor-bearing mice that received injections of an anti-VEGF receptor (VEGFR)-2 monoclonal antibody, such as DC101. Rapid increases of mouse VEGF (e.g., within 24 hours) up to 1 order of magnitude were observed after single injections of DC101 in non-tumor-bearing severe combined immunodeficient or nude mice; similar increases in human plasma VEGF were detected in human tumor-bearing mice. RAFL-1, another anti-VEGFR-2 antibody, also caused a significant increase in plasma VEGF. In contrast, increases in mouse VEGF levels were not seen when small molecule VEGFR-2 inhibitors were tested in normal mice. Most importantly, the increases in plasma VEGF were induced in a dose-dependent manner, with the maximum values peaking when doses previously determined to be optimally therapeutic were used. Plasma VEGF should be considered as a possible surrogate pharmacodynamic marker for determining the optimal biological dose of antibody drugs that block VEGFR-2 (KDR) activity in a clinical setting.     

Determination of the rat tissue partitioning of endotoxin in vitro for physiologically-based pharmacokinetic (PBPK) modeling

The biosynthetically double-labeled lipopolysaccharide (LPS), containing (3)H-labeled on the fatty acyl-chains and (14)C-labeled on the glucosamine of Salmonella enterica serotype typhimurium, was isolated from bacteria grown in proteose peptone-beef extract (PPBE) medium in the presence of labeled precursors; 133 micro Ci/ml of [2-(3)H] acetate sodium salt and 0.167 micro Ci/ml of N-acetyl[D-1-(14)C]glucosamine. The LPS was extracted from the bacteria with 90% phenol/chloroform/petroleum ether, purified and stored in 0.1% (v/v) triethylamine/10 mM Tris HCl at -70 degrees C. Tissue slices and portions of the meninges were prepared and incubated in artificial cerebrospinal fluid (CSF) or Krebs phosphate buffer (Krebs) containing 150 ng/ml LPS with [(3)H] LPS (0.004 micro Ci/ml, sp. act. 28 micro Ci/mg LPS). The tissues were incubated under 95% oxygen/5% carbon dioxide at 37 degrees C with constant agitation until steady-state uptake was reached (60 min). At the end of the incubation period, tissues were processed for radioactivity measurement. The rat tissue partitioning of LPS in artificial CSF for brain and Krebs for other organs was measured by using the ratio of tissue to medium at the steady state in vitro. The following results were obtained from the study: Heart, 0.15; liver, 0.19; spleen, 0.12; kidney, 0.18; stomach, 0.17; small intestine, 0.18; brain stem, 0.10; cerebellum, 0.11; meninges, 0.77; hippocampus, 0.12; hypothalamus, 0.12; frontal cortex, 0.09 and caudate nucleus, 0.10. This information, along with plasma or blood/buffer partition coefficients, is a requisite for constructing a physiologically-based pharmacokinetic (PBPK) model of endotoxins for quantitative risk assessment.     

Amelioration of sulfur mustard skin injury following a topical treatment with a mixture of a steroid and a NSAID

The ability to ameliorate sulfur mustard (HD)-induced oedema by treatment with anti-inflammatory drugs was reported previously after screening four steroids and four non-steroidal anti-inflammatory drugs (NSAIDs) using the mouse ear vesicant model. Following the screening study, one steroid and one NSAID (Adexone and Voltaren) were selected as the most effective, and a mixture of the two was chosen for the present more extensive research. The effect of the combined treatment on clinical, biochemical and histopathological parameters following HD insult was studied. Mice ears were exposed to 0.2 micro l of HD for 10 min to produce a moderate skin injury. Oedema development peaked ca. 48 h following exposure, as determined by weighing ear biopsies. Histological observations at that time exhibited damage to the epidermis and dermis. An increase in prostaglandin E (PGE) was measured in skin homogenates, starting 8 h following exposure and lasting at least up to 48 h post-exposure. A topical treatment using the above anti-inflammatory mixture significantly reduced inflammatory parameters when applied up to 4 h following exposure. These parameters included extent of oedema, levels of PGE, area of clinical damage and extent of cytotoxic injury (vesications and damaged epithelial cells). Thus, a combination of a steroid and NSAID was found to be effective in reducing the intensity of HD skin injury and possibly shortening the time to full recovery. The treatment, however, did not prevent completely the ensuing cytotoxic processes in the epithelial layer.     

Effect of lipopolysaccharide contamination on the attachment of osteoblast-like cells to titanium and titanium alloy in vitro

Failing implants with loss of alveolar bone are associated with gram-negative bacteria that carry lipopolysaccharide (LPS) in the bacterial cell wall. Bony regeneration around these implants is still an unpredictable procedure due to the many clinical factors involved. One important factor is the presence of contaminants such as LPS on the implant surface. The effect of implant-associated LPS on the attachment of bone cells to the implant surface is unknown. This project investigated the effect of LPS on the attachment of osteoblast-like cells (MC3T3-E1) to titanium and titanium alloy surfaces in vitro. We hypothesized that LPS would inhibit bone cell attachment either through loss of cellular attachment sites or alteration of cellular function. Three experimental approaches were used. First, alloy surfaces were exposed to LPS (100 microgram/mL) before the cells were allowed to attach. In the second approach, the cells were exposed to the LPS before they were allowed to attach. Last, the cells were allowed to attach before exposure to LPS. Cellular attachment to implant materials was measured by using a histochemical stain (MTT). The results indicated that LPS presence did not significantly (P > .05) alter osteoblast attachment to titanium or titanium alloy surfaces whether the exposure occurred before or after cellular adherence. It was concluded that LPS did not directly effect the attachment of the MC3T3-E1 osteoblasts to these implant surfaces in vitro. Further research is needed to define the clinical liabilities of LPS during implant placement and maintenance.     

What parents should know about estrogen-like compounds in dental materials

The use of pit and fissure sealants has been reported to increase exposure to xenoestrogens. Because these estrogen-mimics are suspected of having many deleterious effects in animals, and perhaps humans, several types of studies were undertaken by our Biocompatibility Group. We confirmed that bisphenol A (BPA) and bisphenol A dimethacrylate (BPA-DM) have proliferative effects in cells with high levels of estrogen receptors. However, BPA was not detected by our group in American-made sealants, and BPA-DM was detectable in only a few. In addition, the surface layer of the sealant can be treated to reduce the possibility of unpolymerized BPA-DM being left on the tooth. We believe it is important to reassure parents that their children are less likely to be exposed to BPA from sealants than from the ingestion of soft drinks or canned food.     

Paracrine immunotherapy with interleukin-2 and local chemotherapy is synergistic in the treatment of experimental brain tumors

Potent immune responses against malignant brain tumors can be elicited by paracrine intracranial (i.c.) immunotherapy with interleukin (IL)-2. Additionally, i.c. delivery of carmustine via biodegradable polymers has been shown to significantly prolong survival in both animal models and clinical trials. In this study, we show that the combination of paracrine immunotherapy, with nonreplicating genetically engineered tumor cells that produce IL-2, and local delivery of chemotherapy by biodegradable polymers prolongs survival in a synergistic manner in mice challenged intracranially with a lethal murine brain tumor. Animals receiving IL-2-transduced cells and polymers containing 10% 1,3-bis(2-chloroethyl)-1-nitrosourea had significantly improved survival compared with animals receiving IL-2-transduced cells or 10% 1,3-bis(2-chloroethyl)-1-nitrosourea alone. Median survival for the control group was 19 days. Survival in animals receiving IL-2-transduced cells and 1% carboplatin-containing polymers was also significantly improved compared with either therapy alone. Histopathological examination on day 14 of animals receiving combination treatment showed rare degenerating tumor cells. In addition to tissue necrosis surrounding the polymer, a marked inflammatory reaction was observed. In long-term survivors (all animals receiving combination treatment), no tumor was observed and the inflammatory reaction was completely resolved. The brains of animals receiving combination therapy showed both tissue necrosis due to local chemotherapy and strong inflammation due to paracrine immunotherapy. The demonstration of synergy between paracrine IL-2 and local i.c. delivery of antineoplastic drugs is novel and may provide a combined treatment strategy for use against both primary and metastatic i.c. tumors.     

Multiple melanomas after treatment for Hodgkin lymphoma in a non-Dutch p16-Leiden mutation carrier with 2 MC1R high-risk variants

BACKGROUND: A 19-base pair germline deletion in exon 2 of the CDKN2A (cyclin-dependent kinase inhibitor 2A) gene (Leiden mutation) has been detected in Dutch families with familial melanomas. The penetrance of CDKN2A mutations varies widely and is influenced by environmental and unrelated genetic factors such as variants in the MC1R gene. OBSERVATIONS: We describe a 25-year-old German woman who developed 8 invasive melanomas and 6 in situ melanomas after radiation therapy and polychemotherapy for Hodgkin lymphoma. Genetic testing revealed a constitutional CDKN2A Leiden mutation in the proband and her sister, mother, and mother's sister. The proband also carried high-risk MC1R variant alleles R151C and R160W, which she had inherited from her father and her mother, respectively. The less affected mutation carrier sister did not have high-risk MC1R variant alleles. Analysis of DNA from paraffin-embedded tissues showed loss of heterozygosity at CDKN2A loci in all 3 melanomas studied but not in Hodgkin lymphoma. The pedigree revealed several types of cancers on both sides of the family, but no Dutch ancestors were found. No mutations in the CDK4, B-raf, and N-ras genes were detected either in the germline or in tumors from the patient. CONCLUSION: This study shows the variability of the penetrance of the CDKN2A Leiden mutation within the same family, which could be due to genetic or exogenous factors.