Broad epitope coverage of a human in vitro antibody library

Successful discovery of therapeutic antibodies hinges on the identification of appropriate affinity binders targeting a diversity of molecular epitopes presented by the antigen. Antibody campaigns that yield such broad “epitope coverage” increase the likelihood of identifying candidates with the desired biological functions. Accordingly, epitope binning assays are employed in the early discovery stages to partition antibodies into epitope families or “bins” and prioritize leads for further characterization and optimization. The collaborative program described here, which used hen egg white lysozyme (HEL) as a model antigen, combined 3 key capabilities: 1) access to a diverse panel of antibodies selected from a human in vitro antibody library; 2) application of state-of-the-art high-throughput epitope binning; and 3) analysis and interpretation of the epitope binning data with reference to an exhaustive set of published antibody:HEL co-crystal structures. Binning experiments on a large merged panel of antibodies containing clones from the library and the literature revealed that the inferred epitopes for the library clones overlapped with, and extended beyond, the known structural epitopes. Our analysis revealed that nearly the entire solvent-exposed surface of HEL is antigenic, as has been proposed for protein antigens in general. The data further demonstrated that synthetic antibody repertoires provide as wide epitope coverage as those obtained from animal immunizations. The work highlights molecular insights contributed by increasingly higher-throughput binning methods and their broad utility to guide the discovery of therapeutic antibodies representing a diverse set of functional epitopes.     

Outcomes of screening to prevent cancer: analysis of cumulative incidence of cervical abnormality and modelling of cases and deaths prevented

ObjectiveTo determine the frequency of different outcomes in women participating in cervical screening.DesignAnalysis of screening records from 348 419 women, and modelling of cases of cervical cancer and deaths with and without screening.SettingCervical screening programme in Bristol.ResultsFor every 10 000 women screened from 1976 to 1996, 1564 had abnormal cytology, 818 were investigated, and 543 had abnormal histology. One hundred and seventy six had persistent abnormality for two years or more. In the absence of screening 80 women would be expected to develop cancer of the cervix by 2011, of whom 25 would die. With screening 10 of these deaths would be avoided. Comparison of cumulative abnormality rates with numbers expected to develop cancer in the absence of screening suggests that at least 80% of high grade dyskaryosis and of high grade dysplasia would not progress to cancer. The lifetime risk of having abnormal cytology detected could be as high as 40% for women born since 1960.ConclusionsScreening is labour and resource intensive. It involves treatment for many women not destined to develop invasive cancer. The increased intervention rate for cervical abnormality in England is due to change in practice, not a cohort effect, and is probably the reason for the marked fall in incidence and mortality during the 1990s. For other cancers there is scope for major iatrogenic harm from screening because of invasive tests and treatments.

What is already known on this topic

Since the mid-1980s incidence of and mortality from cervical cancer in women born since the 1930s in England and Wales has fallen; screening is the most likely explanationFor each death prevented many women have to be screened and many are treated who would not have developed a problem

What this study adds

In the NHS cervical screening programme around 1000 women need to be screened for 35 years to prevent one deathOver 80% of women with high grade cervical intraepithelial neoplasia will not develop invasive cancer, but all need to be treatedFor each death prevented, over 150 women have an abnormal result, over 80 are referred for investigation, and over 50 have treatmentBefore the 1988 relaunch of screening with strict quality standards, for each death prevented there were 57 000 tests and 1955 women had abnormal results     

LiCl-induced sickness modulates rat gustatory cortical responses

Gustatory cortex (GC), a structure deeply involved in the making of consumption decisions, presumably performs this function by integrating information about taste, experiences, and internal states related to the animal’s health, such as illness. Here, we investigated this assertion, examining whether illness is represented in GC activity, and how this representation impacts taste responses and behavior. We recorded GC single-neuron activity and local field potentials (LFPs) from healthy rats and rats made ill (via LiCl injection). We show (consistent with the extant literature) that the onset of illness-related behaviors arises contemporaneously with alterations in 7 to 12 Hz LFP power at approximately 12 min following injection. This process was accompanied by reductions in single-neuron taste response magnitudes and discriminability, and with enhancements in palatability-relatedness—a result reflecting the collapse of responses toward a simple “good-bad” code visible in the entire sample, but focused on a specific subset of GC neurons. Overall, our data show that a state (illness) that profoundly reduces consumption changes basic properties of the sensory cortical response to tastes, in a manner that can easily explain illness’ impact on consumption.

Sickness is an internal state that impacts consumption, and so could be expected to influence the neural processing of tastes. This study shows that onset of illness changes basic properties of gustatory cortical network processing and taste responses, such that activity comes more purely to reflect the "goodness" or "badness" of tastes.     

Philippine rubber plantations

Because of a local law, limiting land operated by any one source of private capital to 2,500 acres, rubber plantations have not been extensively developed in the Philippines, but there are at present seven plantations within this limitation in the islands.     

Growth of Cephalotaxus harringtonia plant-cell cultures

Summary Cell cultures of Cephalotaxus harringtonia were examined to characterize growth kinetics. The requirement for an undefined medium supplement (coconut water) was eliminated by maintaining high cell concentrations in semicontinuous and batch growth. Sucrose fed to batch-cultured cells was completely hydrolyzed and a diauxic growth pattern was observed corresponding to first glucose and then fructose uptake. Examination of increases in cell concentrations on the basis of fresh and dry weight showed that a substantial lag period existed between the initiation of substrate uptake and increases in cell volume. Specific growth rates were highest during periods of glucose uptake, but cell yields were comparable for the two sugars. In contrast, studies with glucose or fructose as the sole carbon source indicated that cell yields were significantly lower with fructose but specific growth rates were comparable for the two sugars.Offprint requests to: P. J. Westgate     

Thermotropic behavior of dipalmitoylphosphatidylcholine vesicles reconstituted with the glycoprotein of vesicular stomatitis virus

The vesicular stomatitis virus glycoprotein reconstituted into dipalmitoylphosphatidylcholine (DPPC) vesicles exerts a profound effect upon the DPPC gel to liquid-crystalline phase transition. The glycoprotein was reconstituted into DPPC vesicles by octyl glucoside dialysis. The gel to liquid-crystalline phase transition of these vesicles was monitored by differential scanning calorimetry. Vesicles formed in the absence of glycoprotein (600--2100-A diameter) underwent the phase transition at 41.0 degrees C and had an associated enthalpy change of 8.0 +/- 1.6 kcal/mol. Increasing the mole ratio of glycoprotein to DPPC in the vesicles to 0.15 mol % reduced both the transition temperature and the transition enthalpy change. The enthalpy change as a function of the mole percent glycoprotein could be fit to a straight line by a least-squares procedure. Extrapolation of the results to the glycoprotein concentration where the enthalpy change was zero indicated one glycoprotein molecule bound 270 +/- 150 molecules of DPPC.     

Visualization of planar drug intercalations in B-DNA.

A computerized linked-atom modeling system was developed to examine the stereochemical requirements for intercalation of planar drugs into DNA. All classes of conformational possibilities for extending the polynucleotide backbone were examined for their ability to accommodate insertion of a drug into a base-paired region of DNA compatible with adjacent regions of B-DNA while stacking interactions, steric strain and non-bonded interatomic contacts were optimised. One conformation was found which proved superior to all others in ability to satisfy these criteria: an extension of the backbone by characteristic changes in two torsion angles to trans values, plus a change in one sugar puckering to C3'-endo to relieve strain in an adjacent residue. The turn angle distributed over three polynucleotides for this most general mode of intercalation is 90 degrees, equivalent to a helical unwinding of -18 degrees for B-DNA.