Monoclonal antibodies (Mabs) have long been considered a good class of natural drugs, both because they mimic their natural role in the body and because they have no inherent toxicity. Although rodent Mabs are readily generated, their widespread use as therapeutic agents has been hampered because they are recognized as foreign by the patient. Evidently, clinical Mabs should be as human as possible and results with some of the more recently developed chimerized and humanized Mabs are testimony to this. Mabs that are entirely human are now being produced from phage display and transgenic mice. The first fully human Mabs generated by phage display have already entered clinical trials, and together with recent advances in these technologies, may finally realize the full potential of antibodies.

We transferred the complementarity determining regions from a murine monoclonal antibody that neutralizes infection by respiratory syncytial virus (RSV) to a human IgG1 monoclonal antibody. The resulting reshaped human antibody lost affinity for RSV, but an additional alteration to one of the framework regions restored binding affinity and specificity. This second generation reshaped human monoclonal antibody cross-reacted with all clinical isolates of RSV tested and both prevented disease and cured mice even when administered four days after infection. We expect the antibody will prove useful in the management of this major childhood disease.

The human and mouse met protooncogenes encode proteins that have the characteristics of growth factor receptors. Thus, the 1408 amino acid human met protein (Park et al., 1987) can be divided into several putative domains, including an intracellular protein tyrosine kinase (PTK) domain, a transmembrane domain and a 926 amino acid extracellular domain that possesses a cysteine-rich region. An activa-ed form of the met gene that is present in the chemicallytransformed human cell line (MNNG-HOS) was originally detected by its ability to transform NIH3T3 mouse fibroblasts in DNA transfection experiments (Cooper et al., 1984a, b). Activation of met involves a chromosomal rearrangement in which the regions of the met gene encoding the transmembrane and extracellular domain are replaced by a portion of an unrelated gene that has been designated tpr (Park et al., 1986a; Tempest et al., 1986a). The chimaeric gene is transcribed to produce a 5.0 kb hybrid mRNA that is in turn translated to form a fusion protein. DNA sequence analysis of cDNA clones prepared from transcripts of the activated human met gene reveal that all of the met PTK domain is retained in the product of the activated gene and that the region of the fusion protein encoded by the tpr gene exhibits weak homology to laminin Bi (Chan et al., 1987). Alterations of met were also observed in lines of spontaneously transformed mouse fibroblasts where a modest (4-8 fold) amplification of the protooncogene is accompanied by dramatic (50-100 fold) increase in the level of an 8.5 kb met transcript (Cooper et al., 1986).Northern analysis of mRNAs from a series of human cell lines has revealed a complex pattern of transcription of the met protooncogene (Park et al., 1986). Many cell lines, including a human fibroblast cell line, contain a single 9.0 kb mRNA species. Other cell lines such as the CaLu-I lung tumour line contain both 9.0 kb and 7.0 kb mRNAs while the most complex pattern of transcription of the normal met gene is present in MNNG-HOS cells and in the parent HOS cell line, which both contain 9.0 kb, 7.0 kb and 6.0 kb mRNA species. Most B-cell and T-cell tumour lines do not contain detectable levels of met transcripts.Antibodies raised against synthetic peptides corresponding to the carboxyl terminus of the predicted met gene product have been used to detect proteins encoded by the activated and normal met genes (Park et al., 1986b;Tempest et al., 1986b Tempest et al. (1986b), although in this particular study of low level of phosphorylation of a 165 kD protein, which probably corresponds to the 160 kD protein detected by Park et al. (1986b), was also observed.As a first step in determining whether alterations in met can be implicated in the induction of human tumours we have used antipeptide antibodies to examine met protein kinase activity in a series of human tumour cell lines. In addition, to help understand the large quantitative and qualitative variations in met kinase activity observed in these experiments, we have used SDS-polyacrylamide gel electr...

We have identified a class of transformed NIH3T3 mouse fibroblasts that arise at low frequencies in transfection experiments with DNA from both neoplastic and non‐neoplastic cells and that may result from a low level of spontaneous transformation of NIH3T3 cells. DNA from the transformed cells was unable to transform NIH3T3 cells in a second cycle of transfection and, where examined, the cells showed no evidence for the uptake of the transfected DNA sequences. The results of Southern analyses demonstrate that a mouse homologue of the human met oncogene is amplified 4‐ to 8‐fold in 7 of 10 lines of these transformed NIH3T3 mouse fibroblasts. The cells containing the amplified gene also exhibit at least a 20‐fold overexpression of an 8.5‐kb mRNA that is homologous to met. To test the hypothesis that met encodes a growth factor receptor, we examined the binding of platelet‐derived growth factor, epidermal growth factor, insulin‐like growth factor I and gastrin‐releasing peptide to transformed and non‐transformed NIH3T3 cells. The results show that there is no significant elevation of the binding of these growth factors to cells containing amplification and overexpression of met.

We have raised antibodies against a synthetic dodecapeptide corresponding to the carboxyl terminus of the predicted met gene product. Phosphorylation of 60 kDa and 65 kDa proteins on tyrosine residues was observed when immunoprecipitates of cells containing the activated human met gene were incubated with [Y-~~P]ATP. Phosphoproteins with the same molecular masses could be immunoprecipitated from cells metabolically labelled with [32P]orthophosphate. When considered together, these observations indicate that the activated human met gene encodes 60 kDa and 65 kDa proteins that can catalyse autophosphorylation on tyrosine residues.

In this study it is demonstrated that the activated met gene, which was originally detected in the MNNG-HOS chemically transformed human cell line, is a chimeric gene formed by the joining together of two distinct regions of DNA. Rearrangement of cellular DNA in MNNG-HOS cells was demonstrated by Southern analyses, which showed that the MNNG-HOS cell line contained unique met-related DNA fragments that were not detected in the parental cell line, HOS. Chromosomal localization using a series of rodent-human hybrid cell lines showed that the 5' end of the activated met gene is derived from human chromosome 1, in contrast to the 3' end of met which has been previously localized to human chromosome 7. The chimeric gene is transcribed to produce a 5-kb mRNA that is encoded both by regions of the gene derived from chromosome 1 and by regions of the gene derived from chromosome 7. Karyotype analysis of HOS and MNNG-HOS cells has identified several marker chromosomes that involve translocations of chromosomes 1 and 7. The possible location of the activated met locus within these rearranged chromosomes is discussed.

The lethal and mutagenic effects of methyl methanesulphonate (MMS), ethyl methanesulphonate (EMS), and N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) can be dissociated in a mitomycin C (MTC)-sensitive mutant, strain 302, of Micrococcus radiodurans. As regards lethality 302 is extremely sensitive, compared with the wild type, to MTC and decarbamoyl MTC (DCMTC), slightly sensitive to EMS, MNNG, nitrous acid, 7-bromomethylbenz[alpha]anthracene (BrMBA), and N-acetoxy-N-2-acetylaminofluorene (AAAF), and resistant to MMS, hydroxylamine, and ICR 191G. As regards mutability it is, compared to the wild type, very sensitive to MMS, EMS, and MNNG, and slightly sensitive to hydroxylamine and nitrous acid but not to any other agent examined. Alkaline sucrose gradient studies indicate the 302 does not incise DNA containing BrMBA adducts, although it does incise DNA damaged by AAAF but probably not to the same extent as wild type. We put forward the hypothesis that the hypermutability of 302 is due to the non-removal of bases or nucleotides, modified in exocyclic positions, which have altered base-pairing capabilities, while lethality results from the non-removal of bases or nucleotides, also modified in exocyclic positions, which no longer form hydrogen-bonded base pairs.

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