Effects of an EGFR-binding affibody molecule on intracellular signaling pathways.

Effects on intracellular signaling were studied in cells treated with the affibody molecule (ZEGFR:955)2 that targets the epithelial growth factor receptor (EGFR). EGFR is overexpressed in many types of cancers and plays a fundamental role in cell signaling and it is of interest to find targeting agents capable of blocking the receptor. The clinically approved antibody cetuximab (Erbitux) and the natural ligand EGF were included as reference molecules. Two EGFR-rich cell lines, A-431 and U-343, were exposed to the three targeting agents and lysed. The cell lysates were immunoprecipitated with the receptors, or directly separated by SDS-Page. Autophosphorylation of the receptors and phosphorylation of the downstream signaling proteins Erk and Akt, were evaluated by Western blotting. Although the three different agents compete for the same binding site on EGFR, they influenced the signaling differently. The affibody molecule did not induce autophosphorylation of EGFR or any other receptor in the EGFR-family but, in spite of this, induced phosphorylation of Erk in both cell lines and Akt in the A-431 cells. Thus, the results suggest that the signaling pattern induced by (ZEGFR:955)2 is only partly similar to that induced by cetuximab. This makes the affibody molecule a potentially interesting alternative to cetuximab for EGFR-targeted therapy since it might give different therapy-related effects on tumor cells and different side effects on normal tissues.

In vivo and in vitro uptake of 111In, delivered with the affibody molecule (ZEGFR:955)2, in EGFR expressing tumour cells.

The epidermal growth factor receptor, EGFR, is overexpressed in many carcinomas. Targeting this receptor with radionuclides is important for imaging and therapy applications in nuclear medicine. We investigated the in vitro and in vivo properties of a new high affinity EGFR binding affibody molecule, (ZEGFR:955)2, when conjugated with CHX-A''-DTPA and labelled with 111In. The binding time patterns and retention studies were performed using cultured squamous carcinoma A431 cells that overexpress EGFR. In the in vivo studies, female BALB/c nu/nu mice carrying tumours from xenografted A431 cells were used. The in vitro studies showed EGFR specific binding, high uptake and good retention of 111In when delivered as [111In](ZEGFR:955)2. The retention after 72 h of incubation was 38.0+/-1.15% of the initial level. The biodistribution study showed a tumour specific 111In uptake of 3.8+/-1.4% of injected dose per gram tumour tissue 4 h post-injection. The tumour to blood ratio was 9.1 and the tumours could easily be visualized with a gamma camera at this time-point. 111In delivered with [111In](ZEGFR:955)2 gave an EGFR specific uptake and the results indicated that the (ZEGFR:955)2 affibody molecule is a candidate for radionuclide-based tumour imaging. Potential therapy applications are discussed.

Phage display selection of Affibody molecules with specific binding to the extracellular domain of the epidermal growth factor receptor.

Affibody molecules specific for the epidermal growth factor receptor (EGFR) have been selected by phage display technology from a combinatorial protein library based on the 58-residue, protein A-derived Z domain. EGFR is overexpressed in various malignancies and is frequently associated with poor patient prognosis, and the information provided by targeting this receptor could facilitate both patient diagnostics and treatment. Three selected Affibody variants were shown to selectively bind to the extracellular domain of EGFR (EGFR-ECD). Kinetic biosensor analysis revealed that the three monomeric Affibody molecules bound with similar affinity, ranging from 130 to 185 nM. Head-to-tail dimers of the Affibody molecules were compared for their binding to recombinant EGFR-ECD in biosensor analysis and in human epithelial cancer A431 cells. Although the dimeric Affibody variants were found to bind in a range of 25-50 nM affinities in biosensor analysis, they were found to be low nanomolar binders in the cellular assays. Competition assays using radiolabeled Affibody dimers confirmed specific EGFR-binding and demonstrated that the three Affibody molecules competed for the same epitope. Immunofluorescence microscopy demonstrated that the selected Affibody dimers were initially binding to EGFR at the cell surface of A431, and confocal microscopy analysis showed that the Affibody dimers could thereafter be internalized. The potential use of the described Affibody molecules as targeting agents for radionuclide based imaging applications in various carcinomas is discussed.

PATRIC: the VBI PathoSystems Resource Integration Center.

The PathoSystems Resource Integration Center (PATRIC) is one of eight Bioinformatics Resource Centers (BRCs) funded by the National Institute of Allergy and Infection Diseases (NIAID) to create a data and analysis resource for selected NIAID priority pathogens, specifically proteobacteria of the genera Brucella, Rickettsia and Coxiella, and corona-, calici- and lyssaviruses and viruses associated with hepatitis A and E. The goal of the project is to provide a comprehensive bioinformatics resource for these pathogens, including consistently annotated genome, proteome and metabolic pathway data to facilitate research into counter-measures, including drugs, vaccines and diagnostics. The project's curation strategy has three prongs: 'breadth first' beginning with whole-genome and proteome curation using standardized protocols, a 'targeted' approach addressing the specific needs of researchers and an integrative strategy to leverage high-throughput experimental data (e.g. microarrays, proteomics) and literature. The PATRIC infrastructure consists of a relational database, analytical pipelines and a website which supports browsing, querying, data visualization and the ability to download raw and curated data in standard formats. At present, the site warehouses complete sequences for 17 bacterial and 332 viral genomes. The PATRIC website (https://patric.vbi.vt.edu) will continually grow with the addition of data, analysis and functionality over the course of the project.

Improved measurement of /V(cb)/ using B-->D*l nu decays.

We determine the weak coupling /V(cb)/ between the b and c quarks using a sample of 3 x 10(6) BB; events in the CLEO detector at the Cornell Electron Storage Ring. We determine the yield of reconstructed B-->D*l nu; decays as a function of w, the boost of the D* in the B rest frame, and from this we obtain the differential decay rate d Gamma/dw. By extrapolating d Gamma/dw to w=1, the kinematic end point at which the D* is at rest relative to the B, we extract the product /V(cb)/F(1), where F(1) is the form factor at w=1. Combined with theoretical results for F(1) we determine /V(cb)/=0.0469+/-0.0014(stat)+/-0.0020(syst)+/-0.0018(theor).

Improved measurement of Vub with inclusive semileptonic B decays.

We report a new measurement of the Cabibbo-Kobayashi-Maskawa parameter Vub made with a sample of 9.7 x 10(6) BB- events collected with the CLEO II detector. Using heavy quark theory, we combine the observed yield of leptons from semileptonic B decay in the end-point momentum interval 2.2-2.6 GeV/c with recent CLEO II data on B-->X(s)gamma to find Vub = (4.08+/-0.34+/-0.44+/-0.16+/-0.24)x10(-3), where the first two uncertainties are experimental and the last two are from theory.

Observation of exclusive B --> D(*)K(*-) decays.

We report the first observation of the exclusive decays B-->D((*))K(*-), using 9.66 x 10(6) BB pairs collected at the Upsilon(4S) with the CLEO detector. We measure the following branching fractions: B(B--->D(0)K(*-)) = (6.1+/-1.6+/-1.7)x10(-4), B(B(0)-->D(+)K(*-)) = (3.7+/-1.5+/-1.0)x10(-4), B(B(0)-->D(*+)K(*-)) = (3.8+/-1.3+/-0.8)x10(-4), and B(B--->D(*0)K(*-)) = (7.7+/-2.2+/-2.6)x10(-4). The B-->D(*)K(*-) branching ratios are the averages of those corresponding to the 00 and 11 helicity states. The errors shown are statistical and systematic, respectively.

Search for CP violation in tau--> K(pi)nu(tau) decays.

We search and find no evidence for CP violation in tau decays into the K(pi)nu(tau) final state. We provide limits on the imaginary part of the coupling constant Lambda describing a relative contribution of the CP violating processes with respect to the standard model to be -0.172

Minor surgery at hospitals and clinics in a Kenyan district.

BACKGROUND: Minor surgical operations are performed at almost all hospitals, health centres and dispensaries in Africa. A large proportion of the operations are urgent, prompted by injuries and acute infections. They are rarely recorded and reported systematically, and rates and patterns are poorly known. OBJECTIVE: To describe of all minor surgery performed on a rural African population during one year. DESIGN: Prospective recording and analysis of minor surgical procedures and of the patients undergoing the same procedures. SETTING: Hospitals, health centres, dispensaries and mission clinics in Meru district, Kenya. RESULTS: Out of 26,858 minor operations performed (2,066/100,000 people/year) 22,838 were done at the five hospitals in the district while 2,510 were done at 49 mission clinics (4.3/clinic/month), 764 at ten health centres (6.4/health centre/month) and 620 at 29 dispensaries (1.8/dispensary/month). The most common operations were episiotomy, tooth extraction, wound suture and incision and drainage of abscess. More minor surgery is done on women than on men, and the difference is related to obstetrical procedures. CONCLUSIONS: Relatively few operations were performed at health centres, dispensaries and mission clinics. Possible explanations include perceived quality of care, staff motivation, poor service access outside office hours and service charges.

Bounds on the CP asymmetry in like-sign dileptons from B(0)B*(0) meson decays.

We have measured the charge asymmetry in like-sign dilepton yields from B(0)B*(0) meson decays using the CLEO detector at the Cornell Electron Storage Ring. We find a(0)(ll) identical with[N(l(+)l(+))-N(l(-)l(-))]/[N(l(+)l(+))+N(l(-)l(-))] = +0.013+/-0.050+/-0.005. We combine this result with a previous, independent measurement and obtain Re(epsilon(B))/(1+ the absolute value of epsilon(B)(2)) = +0.0035+/-0.0103+/-0.0015 (uncertainties are statistical and systematic, respectively) for the CP impurity parameter, epsilon(B).

Observation of B --> phiK and B --> phiK*.

We have studied two-body charmless hadronic decays of B mesons into the final states straight phiK and phiK(*). Using 9.7 million B&Bmacr; pairs collected with the CLEO II detector, we observe the decays B- --> phiK- and B0--> phiK(*0) with the following branching fractions: B(B--->phiK-) = (5.5(+2.1)(-1.8)+/-0.6)x10(-6) and B(B0--> phiK(*0)) = (11.5(+4.5+1.8)(-3.7-1.7))x10(-6). We also see evidence for the decays B0-->phiK0 and B---> phiK(*-). However, since the statistical significance is not overwhelming for these modes, we determine upper limits of <12.3x10(-6) and <22.5x10(-6) ( 90% confidence level), respectively.

Measurement of the relative branching fraction of upsilon(4S) to charged and neutral B-meson pairs.

We analyze 9.7x10(6) B_B pairs recorded with the CLEO detector to determine the production ratio of charged to neutral B-meson pairs produced at the Upsilon(4S) resonance. We measure the rates for B0-->J/psiK((*)0) and B+-->J/psiK((*)+) decays and use the world-average B-meson lifetime ratio to extract the relative widths f(+-) / f(00) = gamma(Upsilon(4S)-->B+B-) / gamma(Upsilon(4S)-->B0 B-0)) = 1.04+/-0.07(stat)+/-0.04(syst). With the assumption that f(+-)+f(00) = 1, we obtain f(00) = 0.49+/-0.02(stat)+/-0.01(syst) and f(+-) = 0.51+/-0.02(stat)+/-0.01(syst). This production ratio and its uncertainty apply to all exclusive B-meson branching fractions measured at the Upsilon(4S) resonance.

Surgical output in Kibaale district, Uganda.

BACKGROUND: The output of major surgeries in eastern Africa is low and varies considerably between hospitals and regions. This study in rural Uganda relates surgical output to number of hospital beds, number of in-patient admissions, number of new out-patient visits, estimated catchment area population and number of doctors and nurses. This was to facilitate comparison between institutions and areas and to determine surgical resource needs. OBJECTIVES: To describe the quantitative output of surgery in a Ugandan district and to demonstrate the use of standardised indicators for the purpose of comparison, monitoring, resource needs assessment and planning. DATA SOURCES: Retrospective review and analysis of inpatient and outpatient records at Kagadi district hospital, Kibaale district, Western Uganda, the only hospital providing major surgery in the district. STUDY SELECTION: Data on all recorded major and minor surgical operations in the hospital during two complete calendar years, 1996 and 1997. DATA SYNTHESIS: In 1996 and 1997 there were 331 and 309 major operations performed respectively in Kagadi hospital, Kibaale district. This output corresponded to 3.3 and 3.1 per 100 beds; 270 and 270 per 1,000 in-patient admissions; 328 and 267 per year per 10,000 new out-patient visits; 166 and 155 per 100,000 catchment area population; 83 and 77 per doctor; 27.6 and 25.8 per nurse. Minor surgery output at the same hospital in 1996 and 1997 were 185 and 190 respectively. This corresponded to 183 and 164 per 10,000 new outpatient visits and 93 and 95 operations per 100,000 catchment area population. CONCLUSIONS: The output of surgery in the district was very low in comparison with Europe and North America but within the range common in Eastern Africa. For planning and performance review purposes the most useful indicators are surgical output per 100 beds per year; per 1000 inpatient admissions; per doctor and per nurse per year because they measure workload against available resources. Output per 100,000 estimated catchment area population is useful in relating output to need. These indicators are recommended to be incorporated in district and hospital performance reviews and in annual reports.

Household survey of injuries in a Kenyan district.

OBJECTIVE: To determine the pattern and burden of injuries, their causes and action taken in a rural and urban community in Kenya. DESIGN: Household interview survey and focus group discussions. SETTING: Four rural villages and five urban clusters in Kiambu District, Kenya. SUBJECTS: A total of 1,980 members of 200 rural and 230 urban households. RESULTS: The number of reported injuries was 495, corresponding to 300,000 injuries per 100,000 people per year. Most common were cut or piercing (38.4%), followed by fall (16.2%), burn or scald (14.3%), animal bite or kick (10.1%), hit by moving object (5.9%) and road traffic accident (3.6%). Poisoning, sub-mersion/drowning and explosion were uncommon, each below three per cent. Of all reported injuries, 149 (30.1%) sought care from traditional healers, 91 (18.4%) were subject to self-care, 76 (15.4%) obtained service from drug shops, 22 (4.4%) were brought to a health facility for attention and 17 (3.4%) took no action at all. Additional information was obtained through focus group discussions with students, teachers and members of women groups. These generated detailed information about cases of sexual assault within and outside households which had not been captured during the previous household interviews. CONCLUSION: Injuries are very common but most of them are mild, prompting only home care or no action at all. Only one out of 25 injuries were brought to a health facility for attention. Some types of injury, such as domestic violence and sexual assault, are more likely to be captured through focus group discussions than during household interviews. A combination of methods is likely to best reflect the pattern of injury at community level.

Injuries as a public health problem in sub-Saharan Africa: epidemiology and prospects for control.

Injuries are common and on increase in most developing countries, including sub-Saharan Africa. A large proportion of the injuries are caused by road traffic accidents, falls, burns, assaults, bites, stings and other animal-related injuries, poisonings, drownings/near-drownings and suicide. Globally, injuries are responsible for about five per cent of the total mortality, and the overall global annual costs were estimated in the late 1980s at around 500 billion US dollars. The burden and pattern of injuries in Africa and other developing areas are poorly known and not well studied. The incidence is on the increase, partly due to rapid growth of motorised transport and to expansion of industrial production without adequate safety precautions. This is a review of data on various kinds of injuries in developing countries with a focus on sub-Saharan Africa. A computerised search of the relevant literature published between 1985 and 1998 was conducted and a manual search of journals publishing texts on health in low-income countries and in tropical environments was also done. A few studies on injury prevention policy and on research related to injury epidemiology and prevention have also been identified and included. It is concluded that in a relatively typical East African area with a total mortality rate of 1,300/100,000/year, injuries are likely to cause around 100 of these deaths. The corresponding total rate of significant injuries is estimated at 40,000/100,000/year with a breakdown as tabulated below. [table: see text] Although a few surveys and other investigations of injuries have been conducted over the years, injury epidemiology and control remain under-researched and relatively neglected subject areas. Much needs to be done. Collection and analysis of injury data need to be standardised, for example regarding age groups, gender disaggregation and severity. Injuries and accidents should be subdivided in at least road traffic injury, fall, burn, assault, poisoning, drowning, suicide, homicide and others, and details regarding time and place, victim and main cause should be noted. Morbidity survey field staff should be informed that injuries are part of the illness concept and that questions should be asked accordingly. Details regarding the circumstances surrounding different injuries must be known to those who develop preventive programmes. Injury is a public health problem affecting some people more than others. Our ordinary environment--the home, the work-site, the street or road--represents various kinds of risk, and some of these are difficult to eliminate. Not only do we have to accept much of our environment with its existing houses, equipment, vehicles, transport systems, energy supply, toxic substances etcetera, many also suffer from various inherited or acquired conditions that increase the risk. We therefore need to develop safer and more "forgiving" living environments where ordinary people can live and move around safely. Injury control activities may focus on different categories of injury. Road safety measures often include information and education campaigns, improved driver training, road design and maintenance, regular vehicle safety checks, separation of pedestrians from vehicle traffic, speed limits, safety belt, air-bag and helmet use, special training and control of public service vehicle drivers, bicycle lane separation, road lighting, reflectorised materials on clothing, review of the road traffic related legislation and law enforcement, and emergency medical services improvement. Domestic injuries can be prevented for example with window guards, child barriers at stairs, smoke detectors, clothes and furniture in less flammable materials, replacement of open stoves, stabilising of open lamps, fire-fighting equipment and practice, child-proof poison packaging and storage, safe disposal of toxic waste, home safety education of parents, and strict building code enforcement. Occupational injuries can largely be prevented if well adapted to the work environment. Research is required in several areas. An improved facility-based injury recording and reporting system needs to be developed and tested. There is need to combine data collection methods, such as interview surveys, hospital records, police records, focus group discussions and key informant interviews. The outcome of emergency medical care and of different forms of transport and referral needs to be determined. Different combinations of preventive interventions needs to be evaluated. This review is intended as guidance for those who need a broad overview of the subject of injury occurrence and prevention in Africa, for example in preparation for the development of injury control programmes or to help identify issues requiring further research in this field.

Airborne surgical outreach services in eastern Africa.

BACKGROUND: African Medical and Research Foundation (AMREF) has for forty years offered airborne specialist surgeons or surgical teams to supplement the major surgery conducted by local hospital staff, particularly in Kenya and Tanzania. OBJECTIVE: To quantify and qualitatively describe the contribution of airborne surgical outreach services in relation to the total surgical output of remote rural hospitals in Kenya and Tanzania. DATA SOURCES: Surgical theatre registration books of one government and two church/mission owned hospitals in Kenya and Tanzania from 1994 to 1995; and service records kept by the surgeons during outreach trips in the same period. RESULTS: AMREF contributed 15.1% of all major operations, and operations performed by AMREF were relatively complex. The average number of AMREF's major operations performed per visit was 17 and per visit day 4.3. Numerous minor operations and examinations, ward rounds and an average of 10 patient consultations per day were done during the visits. CONCLUSIONS: Total population-based operation rates, and rates by sex and age, are useful indicators for monitoring and comparison. Every hospital should report, annually, major operations performed by type and by age and sex of patients, with special mention of operations and consultations performed by visiting teams, if any. Each hospital should also state the estimated population of its current catchment area.

Socio-cultural information in support of local health planning: conclusions from a survey in rural Kenya.

Existing health information systems in developing countries are managed and used mainly by biomedically trained personnel and by general healthcare administrators. They focus on epidemiology, service utilization and finance; they generate little of the socio-cultural data needed for developing and adjusting health services and disease control programmes to local health-related perceptions, values and resources. This paper reviews some of the published literature on socio-cultural factors related to health and summarizes the findings of a health household interview study in rural Kenya. It concludes that local views on specific disease categories and their causes are crucial for the planning of local disease control and that health information systems need to be expanded to incorporate such data. It discusses methods of generating community-based socio-cultural health information likely to be useful, particularly at district level. Options include observation, household interviews, focus group discussions and key informant interviews. Four improvements are suggested: (1) health information systems should be reoriented to focus less on individual patients and clients and more on population-based information including socio-cultural data; (2) post-basic health manpower training is needed in applied health-related social sciences; (3) staff positions for social scientists at central, provincial and district levels of the health sector; and (4) expansion of research on national and local socio-cultural issues of relevance to health and care-seeking behaviour.

North-South health research collaboration: challenges in institutional interaction.

North-South health development cooperation often includes research financed largely by external donors. The cooperation varies between projects and programmes with regard to subject area, mix of disciplines involved, research methods, training components and project management arrangements. A variety of problems is encountered, but they are rarely described and discussed in published project reports. We authors conducted a study of a small number of European health researchers collaborating with researchers from the Third World. We focused upon projects involving both biomedical and social science researchers, and apart from a literature review three methods were applied: self-administered questionnaires to European researchers, semistructured interviews with five IHCAR researchers, and written summaries by the three authors, each on one recent or ongoing collaborative project of their choice. Most collaborative projects were initiated from the North and are monodisciplinary or partly interdisciplinary in the sense that researchers did independent data collection preceded by joint planning and followed by joint analysis and write-up. There may be disagreements concerning remuneration such as allowances in relation to fieldwork and training. Socio-cultural misunderstanding and conflict was reportedly rare, and no serious problems were reported regarding authorship and publishing. It is concluded that collaborative research is a complex and poorly understood process with considerable potential and worth pursuing despite the problems. Difficulties related to logistics and finance are easily and freely discussed, while there is little evidence that transdisciplinary research is conducted or even discussed. We recommend that published and unpublished reports on collaborative research projects include more detailed accounts of the North-South collaborative arrangements and their management, ethical and financial aspects.