Accuracy of transvaginal ultrasound versus MRI in the PreOperative Diagnostics of low-grade Endometrial Cancer (PODEC) study: a prospective multicentre study.

AIM: To investigate if the diagnostic accuracy of transvaginal ultrasound (TVUS) performed by gynaecologists is sufficient for preoperative assessment of low-grade endometrial cancer (EC) compared to magnetic resonance imaging (MRI). MATERIALS AND METHODS: MRI and TVUS performed by gynaecologists were assessed at the participating centres. The MRI examinations were interpreted by two radiologists at the tertiary centre. Deep myometrial and cervical stroma invasion were visually assessed and compared to postoperative histopathology. RESULTS: Two hundred and fifty-nine patients were included. There was a statistically significant difference in specificity assessing deep myometrial invasion between MRI and TVUS (MRI 0.88, TVUS 0.68). There was no difference in sensitivity (MRI 0.73, TVUS 0.68). When assessing cervical stroma infiltration, MRI had a higher specificity (MRI 0.96, TVUS 0.90), but there was no difference in sensitivity (MRI 0.41, TVUS 0.32). CONCLUSION: MRI has higher specificity than TVUS performed by gynaecologists for assessing deep MI and CSI in low-grade EC, but similar sensitivities. The use of TVUS as a first-line test, rather than MRI, may be supported by this study in centres where access to MRI may be limited.

Evaluation of time, attendance of medical staff, and resources during radiotherapy for breast cancer patients. The DEGRO-QUIRO trial.

BACKGROUND AND PURPOSE: To conform to recommendations regarding the treatment of breast cancer, an estimation of costs and personnel to assure treatment is required. To date no recommendations based on real time measurements are available. The DEGRO (German Society of Radiation Oncology), therefore, initiated a prospective multicenter evaluation of core procedures of radiotherapy. In this analysis, the results regarding human resources and room occupation during the treatment of breast cancer are presented. PATIENTS AND METHODS: Three academic radiation oncology centers (Erlangen, Münster, Mannheim) prospectively documented their workflow and working time for all breast cancer patients from July-October 2008. Subsequently, a statistical analysis was performed. RESULTS: The longest working time of physicians was the definition of the target volume and organs at risk (mean 33 min). Furthermore, physicians needed much time for general tasks, which included conversations. Physicists needed the most time for treatment planning and authorization (64 min), whereas technicians were mostly needed in day-to-day radiotherapy treatment (15 min, 31 min including verification). Despite significant differences in specific steps between centers, overall working times and room occupation were comparable and representative. Special procedures (intraoperative radiotherapy/multicatheter brachytherapy) required considerable amounts of additional working time of physicians and physicists. CONCLUSION: In this prospective analysis, data of human resources and room occupation during treatment of breast cancer are presented for the first time. Each patient consumes about 12 h of human resources for treatment and 3.75 h for general tasks (physicians 4.7 h, physicists 1.8 h, and technicians 9.2 h).

The Euro Heart Survey - Germany: diabetes mellitus remains unrecognized in patients with coronary artery disease.

AIM: Diabetes mellitus is associated with a poor prognosis due to a high rate of coronary artery disease. It was the aim of this survey to assess the prevalence of an impaired glucose tolerance and manifest diabetes mellitus in patients with coronary artery disease (CAD). METHODS: We analyzed data of all German centers participating in the Euro Heart Survey on diabetes and the heart, an European-wide multicenter prospective observational study. Participating centers were asked to recruit patients >18 years with a diagnosis of CAD. RESULTS: In Germany, 261 patients with a diagnosis of CAD were enrolled in five participating centers. Patients were divided into an acutely (22,4%; n = 57) or electively admitted (77,6%; n = 198) group. There were 34% (n = 89) of patients with already known diabetes. In 36% (n = 22 of 56) of the patients without previously known diabetes, an oral glucose tolerance test (OGTT) was performed (3%, n = 5 in the acute and 33%, n = 51 in the elective group). As a result, 39% (n = 22 of 56) of these patients had an impaired glucose tolerance (acute group: 0%, n = 0 of 5; elective group: 43%, n = 22 of 51) and in 13% (n = 7 of 56) diabetes mellitus was diagnosed (acute group: 40%, n = 2 of 5; elective group: 10%, n = 5 of 51). Furthermore, on admission 86% of women and 94% of men reported to exercise less than three times per week and thus less than recommended in current guidelines. CONCLUSION: More than one third of the patients with CAD who underwent an OGTT had an impaired glucose tolerance. Implementation of this simple, effective and inexpensive test into clinical routine of patients with CAD would help diagnose diabetes mellitus and thus grant these high risk patients access to an optimal medical, interventional and surgical therapy. Furthermore, patients ought to be encouraged to include exercise training into their daily routine.

Radiosensitization of tumor-targeted radioimmunotherapy with prolonged topotecan infusion in human breast cancer xenografts.

Clinical radioimmunotherapy (RIT) of solid tumors holds great promise, but as yet has been unable to deliver tumoricidal radiation doses without unacceptable toxicity. Our experimental approach aims to potentiate the therapeutic action of radioimmunoconjugates at the tumor site and thus improve the efficacy of RIT by combination with other treatment modalities. The topoisomerase I inhibitors are a unique class of chemotherapeutic agents that interfere with DNA breakage-reunion by inhibiting the action of topoisomerase I. Preclinical studies suggest that prolonged infusion of topoisomerase I inhibitors enhances cell toxicity due to ionizing radiation. We evaluated the efficacy of combined treatment with continuous administration of topotecan and 90Y-MX-DPTA BrE3 monoclonal antibody (which recognizes an epitope of breast epithelial mucin expressed in most breast cancers) on human mammary carcinoma xenografts in nude mice. Topotecan or 90Y-BrE3 treatment alone delayed overall tumor growth rate transiently but did not affect survival. The combination of RIT with topotecan substantially reduced growth of relatively large established tumors and caused complete tumor regressions and prolonged tumor-free survival in a substantial proportion of treated animals. In vitro studies demonstrated an increase in apoptotic rate and a decrease in cell proliferation of tumor cell lines treated with this combination. We combined the radiosensitization property of topotecan and the specificity of systemic RIT to establish a novel therapy for solid tumors in an experimental tumor xenograft model.

pi bonding in second and third row molecules: testing the strength of Linus's blanket

The flexibility of valence bond (VB) theory provides a new method of calculating pi-bond energies in the double-bonded species H(m)A=BH(n), where A, B = C, N, O, Si, P, S. This new method circumvents the problems usually associated with obtaining pi-bond strengths by targeting only the pi bond, while all other factors remain constant. In this manner, a clean separation between sigma- and pi effects can be achieved which highlights some expected trends in bond strength upon moving from left to right and up and down the Periodic Table. Intra-row pi bonds conform to the classic statement by Pauling [L. Pauling, The Natiure of the Chemical Bond, Cornell University Press, Ithaca, 1960, 3rd edition] regarding the relationship of heteronuclear bond strengths to their homonuclear constituents whereas inter-row pi bonds do not. This variance with Pauling's statement is shown to be due to the constraining effect of the underlying sigma bonds which prevents optimal p(pi)-p(pi) overlap. While Pauling's statement was based on the assumption that the resonance energy (RE) would be large for heteronuclear and small for homonuclear bonds, we have found large REs for all bonds studied herein; this leads to the conclusion that REs are dependent not only on the electronegativity difference but also the electronegativity sum of the constituent atoms. This situation where the bond is neither covalent nor ionic but originates in the covalent-ionic mixing has been termed charge shift (CS) bonding [S. Shaik, P. Maitre, G. Sini, P. C. Hiberty, J. Am. Chem. Soc. 1992, 114, 7861]. We have shown that CS bonding extends beyond single sigma bonds in first row molecules, thus supporting the idea that CS-bonding is a ubiquitous bonding form.

Advance directives in a pediatric emergency department.

Parents of children with terminal illnesses are not always present when a life-threatening event occurs. For many of these children, an advance directive specifying alternate code orders has been written by the parent or legal guardian (hereafter the use of parent is to be interpreted as parent/legal guardian) and the patient's attending physician. Implementing a pediatric advanced directive presents significant problems for emergency personnel if the parent is not present to identify the existence of the directive and interpret the contents in the context of the current medical crisis.

Initial clinical evaluation of radiolabeled MX-DTPA humanized BrE-3 antibody in patients with advanced breast cancer.

To evaluate radiometal-labeled humanized BrE-3 (huBrE-3) monoclonal antibody as a radioimmunolocalization and therapeutic agent in breast cancer patients, tumor localization, pharmacokinetics, radiation dosimetry, and immunogenicity of (111)In-labeled combined 1-p-isothiocyanatobenzyl 3-methyl- and 1-p-isothiocyanatobenzyl 4-methyldiethylenetriamine pentaacetic acid (MX-DTPA) huBrE-3 were studied. Seven women with BrE-3 antigen-positive, metastatic breast carcinoma underwent (111)In huBrE-3 infusion (5 mCi; 50 mg), followed by serial gamma camera imaging and plasma sampling. Region of interest analysis of images was used to make radiation absorbed dose estimates for (111)In huBrE-3. Data were extrapolated to 90Y huBrE-3. Human anti-human antibody (HAHA) response was measured in serum samples obtained up to 3 months after infusion. Patients tolerated infusions well. Seventy-six percent of 105 known sites of disease were identified on planar and single-photon emission computed tomography scans. For six of seven patients, a biexponential model fit the plasma time-activity curve best with an average T1/2alpha=10.6+/-8.5 (SD) h and average T1/2beta=114.2+/-39.2 h. Radiation absorbed dose estimates for (111)In huBrE-3 for whole body averaged 0.53+/-.08 rads/mCi. Dose estimates for 90Y huBrE-3 for marrow averaged 8.4+/-11.9 rads/mCi, and for tumors, 70+/-31.5 rads/mCi. Liver radioactivity uptake averaged 19.7+/-8.8% injected dose at 24 h after infusion, translating into an average radiation absorbed dose 21.1+/-12 rads/90Y mCi administered. Only one of seven patients demonstrated a low level of HAHA response. Although the plasma half-lives are longer and marrow dose higher for radiolabeled huBrE-3 compared with the murine construct, the excellent tumor localization, good tumor dosimetry, and low immunogenicity support the use of 90Y-huBrE-3 antibody for radioimmunotherapy of breast cancer.

Conjugation of interferon alpha to a humanized monoclonal antibody (HuBrE-3vl) enhances the selective localization and antitumor effects of interferon in breast cancer xenografts.

Human mammary carcinoma xenografts (MCF-7) growing in nude mice were treated with natural interferon alpha (n-IFN-alpha) alone or conjugated to a humanized monoclonal antibody (MoAb) anti-breast mucin (HuBrE-3vl) or to irrelevant human IgG1kappa. The IFN and the conjugates were administered as 20 intra-lesional (i.l.) injections to 1 of 2 xenografts in each mouse, or i.p. The growth inhibitory effects of HuBrE-3vl/nIFN-alpha were significantly greater than those of nIFN-alpha used as a single agent or conjugated to HuIgG1kappa. These effects occurred locally in the tumors receiving i.l. injections and systemically, although to a slightly lesser extent, in the noninjected tumors of mice treated i.l. and in the xenografts of mice treated i.p. Biodistribution studies showed that the uptake of 125I-HuBrE-3vl/nIFN-alpha by the tumors 24 hours after i.l. or s.c. injection was greater than that of 125I-HuIgG1kappa/nIFN-alpha, 125I-nIFN-alpha alone, or by normal tissues, documenting a tumor targeting effect and favorable tumor:normal tissues (T:NT) ratios. The targeting effects and the resulting tumor growth inhibition were favored by the IFN-mediated up-regulation of the HuBrE-3vl reactive antigen, which was more prominent after 3 weeks of treatment with HuBrE-3vl/nIFN-alpha. These results were superior to those we obtained previously with nIFN-alpha conjugated to another MoAb of the same group (Mc5). These studies point out the potential usefulness of HuBrE-3vl/nIFN-alpha for the local and systemic treatment of breast cancer lesions by providing a means of delivering high doses of IFN to the tumors while minimizing the amount of IFN binding to normal tissues.

Biodistribution of anti-breast mucin HuBrE3-derived inverted Fabs (IFabs) in nude mice carrying MX-1 xenografts.

In radioimmunotherapy, the long circulation times of antibody radioconjugates correlate with high relative radiation doses to nontumor tissues. Tumor/normal tissue ratios can be significantly improved by using targeting molecules with shorter circulation times. IFabs are multimers of VH-CH1-linker-VK-CK monomers. The lack of the Fc region in IFabs should lead to circulation times that are shorter than those of IgG molecules. The monomers assemble into disulfide-bond-stabilized multimers, 90% of which are 100 kDa dimers (IFab2). IFab2s should not be rapidly eliminated through kidney filtration because their molecular weight is above the threshold for renal passage. We report the first experimental in vivo tests for 125I-IFab radioconjugates derived from a humanized version of the anti-breast mucin monoclonal antibody BrE-3. Biodistributions are reported for athymic nude mice carrying human mammary tumor MX-1 xenografts. The T1/2 beta's for the different tissues ranged from 13.3 h for blood to 19.9 h for tumor. Therefore, IFab radioconjugates cleared the body with a rate comparable to that of F(ab')2 fragments. Except for stomach, tumor/nontumor dose ratios were significantly better for IFabs than for the parent antibody (BrE-3)4 days after injection.

Effect of multiple, repeated doses of radioimmunotherapy on target antigen expression (breast MUC-1 mucin) in breast carcinomas.

The effect of radioimmunotherapy (RIT) on target antigen expression was studied in breast carcinomas transplanted in immunodeficient mice. In nine separate experiments, a single dose of 1500 microCi of 131I-labeled monoclonal antibody (MAb) Mc5 was given to groups of mice carrying well-established, vascularized, transplantable breast tumors (MX-1). Mc5 recognizes an epitope on the tandem repeat of the breast epithelial MUC-1 mucin. This dose suppressed tumor growth for at least 20 days, after which the tumors began to regrow. At various times thereafter, tumors were removed and analyzed for target antigen expression by flow cytometry and immunohistochemistry. In no case was there any significant decrease in antigen content/cell in the tumors of treated mice compared to tumors in control untreated mice. Similar results were obtained with four other breast carcinomas (MCF-7, MDA-MB-331, MDA-MB-435, and MX-2A). To assess the effect of repeated RIT doses on target antigen expression, groups of mice with MX-1 tumors were given 2, 3, and 4 consecutive doses of 1200 microCi of 131I-labeled Mc5. One mouse each at 2, 3, and 4 doses (3 of 18) was cured of its tumor. Control mice were sacrificed after 50 days due to the excessive size of their tumors. Tumors from four mice from each group (2, 3, and 4 doses), after they began to regrow, were excised and analyzed for mucin content and compared to tumors from untreated mice with similar-size tumors transplanted at later dates. In none of the treated groups was there any decrease in mucin content. These results demonstrate that RIT with an anti-breast mucin MAb does not result in the appearance of antigen-negative tumor cells, thus indicating that repeated fractionated doses, which will most likely be necessary for an eventual cure of breast cancer with MAb therapy, are possible.

Extracellular proteinase activity of Cryptococcus neoformans.

Extracellular proteinase activity was studied for eight strains of Cryptococcus neoformans var. neoformans and two strains of Cryptococcus neoformans var. gattii. Proteinase activity was measured by protein agar clearance, azoalbumin hydrolysis, gelatin liquefaction, and protein substrate polyacrylamide gel electrophoresis. All strains of C. neoformans produced extracellular proteolytic activity. Maximal extracellular proteinase activity in supernatants of C. neoformans cultures was associated with late logarithmic- and stationary-phase cultures. C. neoformans was able to utilize murine immunoglobulin G1, bovine immunoglobulin G, and human complement factor 5 for growth in media containing these proteins as the sole sources of carbon and nitrogen, suggesting a capacity to degrade immunologically important proteins. Protein substrate polyacrylamide gel electrophoresis revealed several bands with proteolytic activity at apparent molecular masses of 200, 100, and 50 kDa. The results confirm the existence of extracellular proteinase activity for C. neoformans.

Biological activity of two humanized antibodies against two different breast cancer antigens and comparison to their original murine forms.

We have humanized two monoclonal antibodies (MoAbs), hu-BrE-3 and hu-Mc3, that are bound to two different antigens of the breast epithelial cell. They bind to the breast epithelial mucin (M(r) 400,000) and the BA46 antigen (M(r) 46,000). They could participate in a joint radioimmunotherapy strategy administering repeated or fractionated dosages, where increased irradiation could be delivered by their simultaneous administration. Both antibodies, hu-BrE-3 and hu-Mc3, had similar reactivity to their antigens and similar binding affinity as those of their original murine forms. However, because humanized MoAbs could have different pharmacokinetic and radioimmunotherapeutic characteristics than their original murine forms, the experimental biodistribution in vivo of both of these two humanized anti-breast tumor MoAbs was compared to their original murine forms. Biodistributions in immunodeficient mice grafted with transplantable human breast tumors, both after radioiodination and 111In labeling via 1-p-isothiocyanatobenzyl-methyl-diethylene-triaminepenta-ace tic acid (MXDTPA), demonstrated comparable tumor:normal tissue ratios for the humanized and murine forms. In radioimmunotherapy, the humanized forms for both MoAbs showed also similar tumoricidal activity as that of the original murine MoAbs. These results show that the new humanized forms are amenable to conjugation and radioisotope labeling without loss of biological activity. Furthermore, they demonstrate that these engineered molecules kept intact, both qualitatively and quantitatively, their binding ability, pharmacokinetics, and radioimmunotherapeutic characteristics after the humanization process.

Anti-BA46 monoclonal antibody Mc3: humanization using a novel positional consensus and in vivo and in vitro characterization.

Mc3 is a murine mAb that is highly effective in treating breast tumors in experimental radioimmunotherapy. Mc3 binds to BA46, a 46-kDa glycoprotein of the human milk fat globule membrane that is also expressed in breast carcinoma cells. We cloned and sequenced cDNAs encoding the variable regions of Mc3 and constructed an IgG1, kappa human/mouse chimeric antibody. We then humanized the variable regions of Mc3 using a positional consensus method and retaining residues that might either contact the complementarity-determining regions or the opposite chain. This positional consensus is novel in that it does not include residues with buried side chains. Humanized Mc3 retained full binding affinity, and fully competes with murine Mc3 for antigen binding. Humanized and murine 131I-labeled Mc3 behaved identically in athymic nu/nu mice biodistribution studies. The tumor uptake levels for both antibodies increased over a period of 4 days within a range of 13-20% of the injected dose per g with extremely favorable tumor:normal ratios. Also, a single therapeutic dose of 131I-labeled humanized Mc3 in the same animal model reduced the average tumor size and produced one of five cures while in the uninjected control tumor growth continued unabated. We believe that these results justify the implementation of Phase I human clinical trials for imaging and radioimmunotherapy of breast cancer.

Methods of assessing motility of the digestive system in children.

The study of gastrointestinal motility in children has evolved during the past 25 years. Miniaturization of tools for collecting data has created opportunities to study the maturation of gastrointestinal motility patterns in infants and complaints of abdominal pain, nausea, diarrhea, constipation and distention in children. Available methods, indications for testing, and data evaluation of pediatric esophageal, gastrointestinal, and colonic motility and manometric tests are discussed.

Humanization of KC4G3, an anti-human carcinoma antibody.

We have previously constructed a chimeric version of KC4G3, a murine antibody that reacts with several human epithelial cancers and binds to the human breast epithelial mucin. We have now successfully humanized KC4G3 using positional consensus data, previously compiled after examining several other antibody structures, listing residues in the VH and V kappa frameworks that could influence antigen binding. We have previously showed that a fraction of the kappa chains of murine and chimeric KC4G3 migrates abnormally on SDS-PAGE most likely due to N-linked glycosylation in V kappa. The glycosylation signal has now been removed from V kappa, as a consequence of humanization. As expected, the humanized kappa chain migrates normally on SDS-PAGE. We detected no significant differences either in the affinities (1.6 x 10(9) M-1 vs. 1.4 x 10(9) M-1, respectively) or in the ability to compete for antigen binding, between the murine and the humanized antibodies. The humanized version is an IgG1, kappa immunoglobulin produced by mouse myeloma SP2/0-Ag14 cells and is designated HuKC4v2. The HuKC4v2 frameworks conform to the V kappa II and VHIII human consensus in all but six positions in V kappa and three positions in VH.