John E. Fothergill

Matrix metalloproteinase-1 is associated with poor prognosis in colorectal cancer.

Colorectal cancer is one of the commonest malignant tumors and has a relatively poor prognosis. The outcome depends on the extent of local and particularly metastatic tumor spread. The matrix metalloproteinases (MMPs) are a family of closely related enzymes that degrade the extracellular matrix and are considered to be important in facilitating tumor invasion and spread1–3. Using immunohistochemistry we have investigated the occurrence in colorectal cancer of MMP–1 (interstitial collagenase). Our monoclonal antibody was prepared against a synthetic peptide corresponding to an amino acid sequence specific for MMP–1 and was selected to react in formalin–fixed wax–embedded sections, thus allowing use in diagnostic histopathology and also enabling access to archival material. We found that the presence of MMP–1 in colorectal cancer is associated with a poor prognosis (P = 0.006) and has prognostic value independent of Dukes stage. One MMP inhibitor that strongly inhibits MMP–1 has already been shown to inhibit growth of human colon cancer xenografts in nude mice4. Our results suggest that treatment of those individuals whose colon tumors produce MMP–1 with MMP inhibitors is a therapeutic strategy worth pursuing.

Matrix metalloproteinase‐1 is associated with poor prognosis in oesophageal cancer

The matrix metalloproteinases (MMPs) are a family of closely related proteolytic enzymes which are involved in the degradation of different components of the extracellular matrix. There is increasing evidence to indicate that individual MMPs have an important role in tumour invasion and tumour spread. Monoclonal antibodies specific for MMP‐1, MMP‐2, or MMP‐9 have been produced, using as immunogens peptides selected from the amino acid sequences of individual MMPs. The presence of MMP‐1, MMP‐2, and MMP‐9 in oesophageal cancer was investigated by immunohistochemistry on formalin‐fixed, wax‐embedded sections of oesophageal cancers. The relationship of individual MMPs to prognosis and survival was determined. MMP‐1 was present in 24 per cent of oesophageal cancers, while MMP‐2 and MMP‐9 were present in 78 and 70 per cent of tumours, respectively. The presence of MMP‐1 was associated with a particularly poor prognosis (log rank test 8·46, P<0·004) and was an independent prognostic factor (P=0·02). The identification of individual MMPs in oesophageal cancer provides a rational basis for use in the treatment of oesophageal cancer of MMP inhibitors which are currently undergoing clinical trial.

Spot the differences: proteomics in cancer research

Separation of thousands of cellular proteins by two-dimensional electrophoresis allows the detailed comparison of proteins from normal and diseased tissue. Mass spectrometry provides a fast and reliable way of characterising proteins of interest, particularly when the gene sequence of the source organism is known. The availability of the human genome sequence has opened up the possibility of identifying protein differences between normal and diseased tissue, thus providing the opportunity to search for tumour markers or for therapeutic targets. This new technology will give much-needed insight into the molecular mechanisms of tumour development and progression.

Combining computation with database access in biomolecular computing

Protein structure analysis is a very important application for database technology, with industrial spinoffs. It is also very demanding because sequence search is so very different from 3-D structure, search. We have developed a common data model for integrating sequence and structure data, and for relating different sequence numbering schemes to 3-D structures. We Lave been able to use our high-level functional language Daplex to express queries of both kinds, but using alternative storage schemas to get good performance in a way that is transparent to the user. Daplex functions can be stored in the database and associated with sub-types in an object-oriented fashion. This architecture allows practising scientists to combine complex geometric calculations with data access, which they need in order to search for complex relationships in the data which may validate or modify their hypotheses.

A simple enzyme-linked immunosorbent assay (ELISA) for the neuron-specific γ isozyme of human enolase (NSE) using monoclonal antibodies raised against synthetic peptides corresponding to isozyme sequence differences

Monoclonal antibodies specific for the gamma isozyme of human enolase (known as neuron-specific enolase or NSE) have been raised against synthetic peptides after coupling to carrier protein: the selected peptides were those corresponding to regions of amino acid sequence difference between the alpha and gamma subunits of these closely similar isozymes. This technique gave monoclonal antibodies of high specificity and affinity. Two monoclonal antibodies raised against different peptides were used to develop a double-antibody sandwich enzyme-linked immunosorbent assay (ELISA), using one as the solid-phase antibody and the other conjugated to horseradish peroxidase to detect the bound NSE. This assay provides a simple and routine method of detecting NSE in serum samples from patients with small-cell carcinoma of the lung and related tumours.

Design of a 3D User Interface to a Database

One of the strangest paradoxes of the silicon era is the dichotomy between ’enjoyable’ recreational computer activities and ’mundane’ work-based computer operations. How can an activity as pointless as a computer game have so much appeal? The answer to this lies in the user interface, and not the functionality, of the program. Computer games rely heavily on an interface which is natural and enjoyable to use. We believe that an interface should appeal to the user, and to do so must capture the users interest and imagination. To this end, we have been using high performance graphics to generate meaningful three dimensional representations for our graphical user interface. We propose new metaphors for both query construction and result representation.

Evidence for a new cytochrome P450 form induced by 3-methylcholanthrene in rats

Evidence is presented for a new 3-methylcholanthrene (3MC)-induced form of cytochrome P450, P450MCX, in rat liver microsomes. P450MCX was co-purified with CYP1A1 from 3MC-treated male Sprague-Dawley rats but was resolved by gel electrophoresis. The M(r) of P450MCX (56,700) was intermediate between CYP1A1 (57,000) and CYP1A2 (54,800). Monoclonal antibodies showed that P450MCX was immunorelated to both CYP1A1 and CYP1A2 but not to CYP2B1, CYP2C6 or CYP3A1. Immunoreactive P450MCX was not detectable in liver microsomes from untreated rats but was highly induced by 3MC and Aroclor 1254, although not induced by isosafrole. The N-terminal amino acid sequence of P450MCX, obtained from an electroblotted sample resolved on SDS-PAGE, did not match any known cytochrome P450 or other protein. P450MCX may be a new member of the CYP1 family.