British Journal of Cancer

Ijiri, K; Potten, C S

doi: 10.1038/bjc.1983.25pmid: 6824565

The spacial distribution of cell death among the epithelial cells lining the adult mammalian small intestinal mucosa at various times after a range of doses of 10 different drugs as well as after internal or external irradiation (beta particles from tritium, gamma- and X-rays and neutrons) has been recorded. Cell death, expressed as pycnosis or apoptosis, has been recorded for each cell position up the side of the crypts of the small intestine. The results, in the form of distributions of dead cells at each cell position, show that each of the various cytotoxic agents tends to act preferentially over a characteristic small range of cell positions. Since cell position is likely to be related to hierarchical cell position within a family tree or cell lineage, each agent tends to act with greatest efficiency on cells at a particular position within the lineage. Adriamycin and the various forms of radiation tend to kill cells preferentially at cell position 4-5 i.e. on cells very early in the lineage, probably stem cells. Isopropyl-methane-sulphonate, nitrogen mustard and possibly Actinomycin-D act on cell position 6-7, while 5-fluorouracil, Myleran, cyclophosphamide, and cycloheximide tend to kill cells at cell position 7-9. Vincristine and hydroxyurea are the 2 agents that exhibit a specificity for cells highest up the crypt, i.e. latest in transit population of the cell lineage by acting on cell positions 10 or 11. The data also suggest that normal healthy cells continue to migrate up the crypt and onto the villus in spite of considerable cell death and reduced cell production.

Twentyman, P R; Workman, P

doi: 10.1038/bjc.1983.26pmid: 6824566

Experiments have been carried out both in vitro and in vivo to examine the possibility of chemosensitization by misonidazole (MISO) at concentrations which are achievable in the clinic. Using multicellular tumour spheroids in vitro we found that a 16 h pre-incubation with 100 micrograms ml-1 MISO under hypoxic conditions led to a considerable enhancement of sensitivity to melphalan (MEL) but not to CCNU. Pre-incubation for 16 h under hypoxia alone also produced a degree of sensitization to MEL, but there was no effect of oxic pre-incubation with MISO. in vivo experiments using the KHT or RIF-1 tumours in C3H mice were designed so that repeated administration of MISO maintained blood concentrations of around 100 micrograms ml-1 for either 7 h or 16 h. For the 7 h regime, cytotoxic drugs were administered at the 4 h point. In most experiments the tumour response to MEL, cyclosphosphamide (CTX), chlorambucil or CCNU was no greater in mice receiving multiple MISO than in mice receiving multiple injections of a balanced salt solution. In the occasional experiment where there was an apparent increase in response, the effect was only small (dose modifying factor less than 1.5). For the 16 h regime the effect was studied of administering CTX (100 mg kg-1) at various times during the regime. There was a clear trend towards increased CTX response in mice receiving multiple MISO compared with controls. There was, however, no clear tendency for the effect to increase with length of MISO pre-exposure.

Murray, D; Meyn, R E

doi: 10.1038/bjc.1983.27pmid: 6824567

The technique of alkaline elution has been adapted for the study of drug-induced DNA cross-link formation in vivo. Pretreatment with misonidazole (MISO) enhances the number of cross-links formed in a fibrosarcoma and in the spleen and gut of mice for periods up to 48 h following a single injection of melphalan (MEL). The tumour was sensitized by a greater factor (2.05) than either of the normal tissues (enhancement factor 1.4-1.5). This enhancement did not appear to be related to inhibition of the repair of actual cross-links. Rather, the effect was explicable in terms of one of two alternative models. Firstly, MISO pretreatment could result in a greater amount of binding of MEL to DNA at early times after injection. This may be the result of altered pharmacokinetics of MEL, or of enhanced intracellular uptake of MEL due to MISO pretreatment. Secondly, MISO may exert its affect by inhibition of the repair of cross-links or monoadducts at early times post-injection, which would not be observed in this study. The possible involvement of glutathione depletion in chemosensitization by MISO was investigated by comparison with the effect of diethyl maleate (DEM), a known thiol-depleting reagent. Glutathione depletion, while perhaps being important, could not account for all of the effects observed.

Morgan, D; Freshney, R I; Darling, J L; Thomas, D G; Celik, F

doi: 10.1038/bjc.1983.28pmid: 6297528

A method has been developed for measuring the drug sensitivity of human gliomas in short-term culture, using scintillation counting or autofluorography. Cell cultures prepared from malignant astrocytomas were treated with anticancer drugs whilst in exponential growth in microtitration plates. After drug treatment and a recovery period, residual viability was measured by [3H] leucine incorporation followed by scintillation counting or by [35S] methionine incorporation and autofluorography in situ. In 5 glioma cell lines tested against 6 drugs, the microtitration method correlated well with monolayer cloning. Although replicate samples of the same tumour showed little variation in chemosensitivity, there was marked variation between the chemosensitivities of cultures derived from the tumours of different patients. However, as variability between replicates was apparent during drug exposure or shortly after, it is important to allow the assay to run as long as possible after drug removal. It is hoped that this assay may provide the basis of a method for the prediction of in vivo chemosensitivity or the screening of potential chemotherapeutic drugs.

Evans, B D; Smith, I E; Millar, J L

doi: 10.1038/bjc.1983.29pmid: 6297529

Immunodeprived mice survived a high, otherwise lethal dose of cyclophosphamide (Cy) provided they had been "primed" with a low dose (50 mg kg-1) of the drug 4 days earlier. These combinations were then tested on 2 oat cell xenograft lines (which are known to reproduce the chemotherapeutic responses of the parent tumours) grown in immunodeprived mice. In the treatment of the first oat cell xenograft, 200 mg kg-1 Cy produced a growth delay of 34 days in the unprimed group and 45 days in the primed group. At a dose of 300 mg kg-1 a growth delay could not be assessed in the control group as 16/17 of these unprimed mice bearing this xenograft died. However, 14/22 tumours went into complete remission in this group before death occurred. In contrast only 3/16 deaths occurred in the group of mice that were primed before receiving the same challenge dose. In these animals 19/26 tumours went into complete remission and were still completely absent when the experiment was terminated at 60 days. Using the second oat cell xenograft, 300 mg kg-1 Cy produced a growth delay of 27 days. However, at this dose level all the animals were dead by day 46. In mice which had been primed with 50 mg kg-1 Cy 4 days before the administration of 300 mg kg-1 a growth delay of 32 days was achieved and 2/9 animals were alive at day 60. This study shows that priming allows larger doses of Cy to be given to immunodeprived mice bearing human tumour xenografts than would normally be tolerated and that the priming does not alter the anti-tumour efficacy of the large challenge dose as measured by tumour growth delay or complete remission rate. As the tumours were human in origin it raises the question whether high dose cyclophosphamide therapy and priming have a role to play in the treatment of patients with oat cell carcinoma.

Silvestrini, B; Hahn, G M; Cioli, V; De Martino, C

doi: 10.1038/bjc.1983.30pmid: 6824568

Lonidamine or 1-[(2, 4-dichlorophenyl) methyl]-1H-indazole-3-carboxylic acid, studied in a battery of in vitro and in vivo tests currently used for the screening of anti-tumour agents affecting cell division, has been shown to have a narrow spectrum of anti-tumour activity. The significance of this finding is discussed in the light of previous investigations suggesting that lonidamine affects mitochondrial function and not cell replication. Hyperthermia has been shown to sensitize tumour cells to lonidamine. This observation indicates that in combination with hyperthermia lonidamine has some potential for the treatment of cancer, moreover, it suggests that hyperthermia might reproduce a metabolic condition occurring in some stages of the disease. The blood levels corresponding to the anti-tumour action of lonidamine in animals are in the range of those detected in patients treated with the drug.

Hastings, R J; Franks, L M

doi: 10.1038/bjc.1983.31pmid: 6572066

To study heterogeneity in a cell line derived from a human bladder carcinoma (EJ), 7 clones were isolated at low passage and examined for differences in culture behaviour, ability to grow in agar and tumorigenicity in nude mice. The parent EJ line had several distinct chromosome populations (both diploid and tetraploid), grew in agar and produced tumours in nude mice. Three of the clones had pseudodiploid modes and 4 had either hypo- or hypertetraploid modes. The 7 clones had 5 marker chromosomes in common but the combination of other marker chromosomes made each clone unique. No significant difference was found between the clones in the in vitro growth rate although analysis of in vitro culture behaviour showed heterogeneity in the pattern of cell movement on plastic substratum. Three clones were composed of static cells, one clone had very mobile cells, the other clones had rates of movement intermediate between the two. Differences were also found in the packing density of the cloned cells and in the cell size. All 7 clones grew in agar but heterogeneity was seen between the clones as shown by widely varying colony-forming efficiencies (0.5-13%). One clone had a high colony-forming ability in agar but failed to produce tumours in nude mice. The other clones were tumorigenic regardless of colony-forming efficiency in agar. Specific chromosome abnormalities were found to be associated with growth in agar and tumorigenicity but not with the growth pattern or the rate of movement of the cloned cells in culture.

Kerr, K M; Robertson, A M; Lamb, D

doi: 10.1038/bjc.1983.32pmid: 6824569

The in vitro thymidine labelling indices (TLI) of 58 human lung tumours were assessed using autoradiography. The labelling technique involved incubation of 1 mm3 tumour fragments with 3H-thymidine (5 muCi ml-1) under conditions of hyperbaric oxygenation at a pressure of 3 atmospheres. Only a rim of labelling was achieved along the edges of fragments and the depth of this rim varied from tumour to tumour. A technique for counting TLIs was therefore devised to take this into account. In general, those tumours showing low TLI values of less than 5.0% showed a greater depth of labelling. The common malignant tumours of the bronchus showed a wide range of values (2.2-30.4%) though the adenocarcinomata had a lower average value than the other groups. With the squamous carcinomata a relationship with differentiation was shown. The mean value for small cell carcinomata (16.9%)--a highly aggressive tumour--was no higher than for the other groups. The low grade malignant tumours showed TLIs of less than 3.0% and these values correlate with their less aggressive clinical behaviour. Labelling of stromal cells and inflammatory cells varied greatly from tumour to tumour, however, no correlation was found with the TLIs of tumour cells.

Smedley, H M; Finan, P; Lennox, E S; Ritson, A; Takei, F; Wraight, P; Sikora, K

doi: 10.1038/bjc.1983.33pmid: 6337613

Rat monoclonal antibodies were prepared by immunising rats with human colorectal carcinoma cell membranes and fusing splenic lymphocytes with a rat myeloma. Hybridoma supernatants were screened by binding assays on membranes prepared from colorectal carcinoma tissue. One hybridoma supernatant, containing a monoclonal antibody with high binding activity on malignant compared to normal colon sections, was grown in large quantities in serum-free medium. After ammonium sulphate precipitation the antibody was purified by ion-exchange chromatography and labelled with 131I. Radiolabelled antibody was administered i.v. to 27 patients with colonic and other tumours. Scintigrams were obtained at 48 h. Computerised subtraction of the blood pool image revealed localised areas of uptake corresponding with areas of known disease in 13/16 patients with colorectal carcinoma and 3/4 patients with breast cancer.

Castagnetta, L; Lo Casto, M; Mercadante, T; Polito, L; Cowan, S; Leake, R E

doi: 10.1038/bjc.1983.34pmid: 6824570

Soluble and nuclear oestrogen receptor status was determined in both the central and peripheral portions of tumour for 37 cases of adenocarcinoma of the endometrium. Of these, 29 had functional receptor in the peripheral biopsy, but only 19 retained functional receptor in the centre. Six of the 10 patients whose tumours showed this difference came from the group of 12 patients who were immediately post-menopausal (4.50 +/- 1.45 y post-menopausal age). Receptor status was not related to tumour classification into histological grades I and II. However, receptor-negative central biopsies were significantly more likely (P less than 0.05) to be Grade III. Early relapse was also related to a receptor-negative central biopsy.