Both PLED and PLEC were shown to exert significant activity against head and neck xenograft tumours, with PLED showing particular efficacy.

A truncated epidermal growth factor receptor (EGFR) expressed from a rearranged and amplified EGFR gene is present at high frequency in gliomas. In this work we show that when this receptor is expressed in NIH3T3 fibroblasts it is partially activated and confers tumorigenicity to this cell line in vivo but no growth advantage in in vitro anchorage-independent growth assays. Because the mutation occurs in the extracellular domain of the receptor, it can be considered to represent a glioma-specific tumour marker. Here we demonstrate that 2 monoclonal antibodies, DH1.1 and DH8.3, raised to a synthetic peptide spanning the unique junctional sequence, can recognise the mutant receptor but not the normal receptor in both denatured and native states. Furthermore, radiolabelled antibody DH8.3 successfully targets tumours expressing this antigen in nude mice.

To determine the potential prognostic value of epithelial cadherin (E-cadherin), a Ca(2+)-dependent cell-cell adhesion molecule, we have analysed its immunoreactivity and cellular localisation in 67 transitional cell carcinomas (TCC) using an avidin-biotin immunoperoxidase technique on formalin-fixed, paraffin-embedded tissues. These results were correlated with histopathological grade, tumour stage, presence of metastases and survival. In addition, 10 cystitis and 11 normal bladder biopsies were evaluated as controls. E-cadherin was expressed in a normal membranous pattern in all normal and 7 of 10 cystitis biopsies. Loss of normal surface E-cadherin expression was found in 3 of 15 superficial tumours and in 48 of 52 invasive cancers. Abnormal immunoreactivity was strictly related to tumour differentiation and stage. Fifteen of 20 well-differentiated (grade I) tumours showed preserved membranous E-cadherin immunoreactivity, while 46 of 47 moderate and poorly differentiated tumours (grades II and III) demonstrated abnormal staining patterns. Loss of membranous E-cadherin immunoreactivity was also associated with advanced tumour stage. There was a significantly higher 5-year survival rate for patients with preserved membranous staining compared with patients with abnormal staining.

The biodistribution and pharmacokinetics of111In-DTPA-labelled pegylated liposomes in tumour-bearing nude mice was studied to examine possible applications of pegylated liposome-targeted anti-cancer therapies. Nude mice received an intravenous injection of 100 μl of111In-DTPA-labelled pegylated liposomes, containing 0.37–0.74 MBq of activity. The t 1/2α and t 1/2β of111In-DTPA-labelled pegylated liposomes were 1.1 and 10.3 h, respectively. Tumour uptake was maximal at 24 h at 5.5 ± 3.0% ID g–1. Significant reticuloendothelial system uptake was demonstrated with 19.3 ± 2.8 and 18.8 ± 4.2% ID g–1at 24 h in the liver and spleen, respectively. Other sites of appreciable deposition were the kidney, skin, female reproductive tract and to a lesser extent the gastrointestinal tract. There was no indication of cumulative deposition of pegylated liposomes in the lung, central nervous system, musculoskeletal system, heart or adrenal glands. In contrast, the t 1/2α and t 1/2β of unencapsulated111In-DTPA were 5 min and 1.1 h, respectively, with no evidence of accumulation in tumour or normal tissues. Incubation of111In-DTPA-labelled pegylated liposomes in human serum for up to 10 days confirmed that they are very stable, with only minor leakage of their contents. The potential applications of pegylated liposomes in the arena of targeted therapy of solid cancers are discussed. © 2000 Cancer Research Campaign

SummaryThe relationship between tumour size and uptake of 111 In-DTPA-labelled pegylated liposomes has been examined in a human head and neck cancer xenograft model in nude mice. The mean tumour uptake of 111 In-labelled pegylated liposomes at 24 hours was 7.2 ± 6.6% ID/g. Liposome uptake for tumours < 0.1 g, 0.1-1.0 g and > 1.0 g was 15.1 ± 10.8, 5.9 ± 2.2 and 3.0 ± 1.3% ID/g, respectively. An inverse correlation between tumour weight and liposome uptake was observed by both Spearman's rank correlation test (r s = -0.573, P < 0.001) and Pearson's correlation coefficient (r s = -0.555, P < 0.001). For 18 tumours with macroscopic central necrosis, the ratio of uptake in the tumour rim relative to the necrotic tumour core was 11.2 ± 6.4. Measurement of tumour vascular volume for tumours of various sizes revealed an inverse correlation between tumour weight and tumour vascular volume (Spearman's rank correlation test, r s = -0.598, P < 0.001), consistent with poor or heterogeneous vascularization of larger tumours. These data have important implications for the clinical application of pegylated liposome targeted strategies for solid cancers which are discussed in detail.

We describe a novel version of antibody‐directed enzyme prodrug therapy (ADEPT), with the use of amygdalin as prodrug. Amygdalin is a naturally occurring cyanogenic glycoside, which can be cleaved by sweet almond β‐glucosidase to yield free cyanide. If amygdalin could be activated specifically at the tumour site, then malignant cells would be killed without the systemic toxicity usually associated with chemotherapy. To this end, we have conjugated β‐glucosidase to a tumour‐associated monoclonal antibody (MAb) (HMFG1) and the conjugate has been tested in vitro for specificity and cytotoxicity subsequent to activation of amygdalin. Amygdalin was cytotoxic to HT1376 bladder cancer cells only at high concentrations, whereas the combination of amygdalin with HMFG1‐β‐glucosidase enhanced the cytotoxic effect of amygdalin by 36‐fold. When 2 concentrations of HMFG1‐β‐glucosidase were compared, the toxic effect was dose dependent. The cytotoxicity of amygdalin was also enhanced by the MAb–enzyme conjugate even when the unbound conjugate was removed from the medium prior to exposure to amygdalin and the cells were washed. In addition to the cytotoxic effect, we also demonstrated specificity, using a MAb–enzyme conjugate that does not recognise the HT1376 bladder cancer cells. Finally, we studied the cytotoxic effect of the conjugate in co‐culture of HMFG1‐positive and ‐negative cell lines (HT1376 and U87MG cells). We demonstrated that the rate of surviving cells corresponds well to the percentage of U87MG (HMFG1‐negative) cells in the flask. Our findings indicate that ADEPT is more effective than non‐directed enzyme activation of a prodrug and can result in a non‐toxic cancer therapy. Int. J. Cancer 78:712–719, 1998. © 1998 Wiley‐Liss, Inc.

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Although much progress has been made in the production of recombinant antibodies and their fusions, there are still problems with solubility and folding. Useful antibodies produced from cloned hybridomas do not always result in scFvs behaving favourably. We report here further work on an scFv (H17E2) against the oncofetal antigen human placental alkaline phosphatase. The overall expression was greatly improved and the H17E2 scFv was redesigned by manipulation of the interdomain linker, resulting in much higher expression levels of the soluble scFv in its active conformation at 0.2-0.5 mg/l of bacterial culture. We show that the new soluble version of this scFv has similar characteristics to the refolded version in terms of antigen and tumour cell binding, stability and in vivo pharmacokinetics. The final tumour uptake behaviour of these scFvs is superior to that of the parental whole antibody with respect to tumour:organ ratios, but still requires further development before considering it as a suitable molecule for clinical use in ovarian or testicular cancer.