Detection of synergistic factor and interleukin-3 activity in the serum and ascites fluid of mice bearing the WEHI-3 tumour

Media conditioned (CM) by WEHI-3 cells (a myelomonocytic leukemia cell line) contains a number of haemopoietic growth factors, including synergistic factor (SF) and interleukin-3 (IL3). We have investigated the production of SF and IL3 in vivo in mice bearing the WEHI-3 tumour. SF and IL3 activity were detected in both the sera and ascites fluids of these mice. SF from the ascites fluid was partially purified by a four-step purification schedule consisting of ammonium sulphate fractionation, DEAE-cellulose, hydroxylapatite, and Sephadex G-100 chromatography. This purification sequence resulted in approximately a 250- and 187-fold purification of SF and IL3 respectively on the initial starting material with a yield of 13 and 9.7% respectively of the initial activity. At each stage of purification, the fractions containing SF co-purified with IL3 activity, further supporting our previous report that SF and IL3 are probably identical molecules. The characteristics of the in-vivo derived (sera and ascites fluid) activities were found to be similar to those of the factors produced in vitro in WEHI-3 cell conditioned media. These results support the conclusion that SF and IL3 are produced in vivo in WEHI-3 tumour bearing mice and are not in vitro artifacts.     

Isolation and identification of a tumour reducing component from mistletoe extract (Iscador)

Using a combination of gel filtration and paper chromatography, a tumour reducing component from mistletoe extract (Iscador) was isolated and identified to be a peptide of approximate molecular weight 5000. The isolated peptide reduced the solid tumour induced by Dalton's lymphoma ascites tumour cells (DLA cells) in mice. The isolated component was very cytotoxic to the DLA cells but was not cytotoxic to normal lymphocytes, indicating a cell dependent specificity.     

Enhanced urinary excretion of 7,8-dihydro-6-hydroxylumazine by mice bearing the Ehrlich ascites tumour.

A fluorescent urinary metabolite, excreted by mice carrying the Ehrlich ascites tumour (EAT), has been identified as 7,8-dihydro-6-hydroxylumazine. This finding agrees with other data which show that a disorder in pteridine metabolism is commonly associated with the presence of malignant growth.     

Effect of purified glutaminase from human ascites fluid on experimental tumor bearing mice

Glutamine is the major respiratory fuel and energy source of the rapidly proliferating tumor cells and that is why glutamine clearance by glutaminase therapy provides an opportunity to fight against the neoplasm. Glutaminase from bacterial source was tried on experimental models but had to be excluded because of its limited efficacy. Search for a better glutaminase continued exploiting the mammalian sources. In the present study, glutaminase purified from human ovarian cancer ascites fluid was used in experimental solid and ascites mice model alone and in combination with Cu-Sulphate and heparin. Cumulative findings indicate that the enzyme alone is quite effective in lowering tumor burden and reducing not only the tumor induced angiogenesis, but also an angiogenic inducer, heparin mediated angiogenesis. However, the presence of Cu with the enzyme, amplified the antineoplastic response by improving anti-angiogenic potential and hematological status of the tumor bearing host. Therefore, Cu-glutaminase combination strengthened the hypothesis that together they may provide a better therapeutic regimen in experimental mice tumor model.     

Investigation of the procoagulant activity present in ascitic fluid and serum of mice bearing the Landschütz ascites carcinoma

The direct procoagulant activity (PCA) of murine tumour cells was found to be more than three orders of magnitude greater than an equivalent concentration of either resident or Corynebacterium parvum-elicited, exudate peritoneal cells. Similarly, a soluble PCA was detected in the extracellular culture medium of only the tumour cells. Studies on the procoagulant nature of the serum and ascitic fluid of tumour-bearing animals suggested that the ascitic fluid may contain a unique PCA factor(s). This activity could not, however, be resolved from inherent procoagulant factors, either by gel filtration or ammonium sulphate fractionation, and a more specific assay for, say, a single enzymic reaction in the coagulation cascade would be required to identify the tumour-associated activity.     

Early systemic effects on the hepatic mitochondria of tumour bearing mice

The activities of citrate synthase and cytochrome c oxidase, mitochondrial marker enzymes, were evaluated in the liver of Ehrlich ascites carcinoma-bearing mice during the life span of the inoculated animals. After 10 days of tumour transplantation, when cell proliferation has ceased, a 30-40% decrease of these activities was detected in both liver and kidney. Simultaneously, an increase in the total acidic proteinase activity (50-60%) was observed. The gel filtration profiles of liver proteinase activities from inoculated animal extracts displayed different patterns to those of the controls; low molecular weight proteinase activities appear to be enhanced in the livers of tumour-bearing animals.     

Tumour enhancement associated with induction of abscesses in mice bearing a transplantable solid tumour

A model of intraabdominal sepsis in the tumour-bearing host was established in order to study the interactions between host, tumour and infecting organisms. BALB/c mice bearing a transplanted tumour were given an intraperitoneal inoculum containing Bacteroides fragilis, Escherichia coli and bran as an abscess-potentiating agent. Tumour-bearing mice formed abscesses which were not significantly smaller than in controls except late in tumour growth. The bacterial contents of the abscesses were not significantly different to controls. In contrast, mice given an abscess-inducing mixture at or near the time of tumour cell inoculation had tumours which were significantly larger than in controls. The mechanism of tumour enhancement is not known.     

Scheduling of gemcitabine and cisplatin in Lewis Lung tumour bearing mice

We used the gemcitabine (dFdC) and cisplatin (cis-diamine dichloroplatinum CDDP) resistant murine NSCLC tumour Lewis Lung (LL) in C57/Bl6 mice to optimise scheduling of both drugs, since in previous in vivo studies no effective combination schedule of both compounds was found to overcome resistance to either drug. dFdC could not be combined at the previously determined maximum tolerated dose (MTD) (120 mg/kg, q3dx4) with CDDP at its MTD (9 mg/kg, q6dx2) (mean weight loss <15% and <15% toxic deaths), because of additive toxicity. Therefore, we lowered the dose of dFdC to 60 mg/kg (q3dx4) and of CDDP to 3 mg/kg (q6dx2), which caused an increase in antitumour effect compared with the activity of each compound alone at its MTD (growth delay factor (GDF)=0.55, 0.13 and 2.56 for dFdC and CDDP alone and the combination, respectively). Changing the CDDP treatment schedule giving the total dose (6 mg/kg) only at day 0 caused unacceptable toxicity. This effect was not seen when mice were treated with the total dose of CDDP on day 9, but, the anti-tumour effect was not enhanced. To decrease toxicity, the dosage of dFdC was lowered to 50 mg/kg and combined with the total dose of CDDP on day 0, which caused a better antitumour effect than the combination of 60 mg/kg dFdC and 3 mg/kg CDDP (q6dx2) with acceptable toxicity. Schedule dependency was found for the combination: dFdC preceding CDDP by 4 h was the best treatment schedule in the LL tumours (GDF: 2.1) with acceptable toxicity. However, when the interval was increased to 24 h, toxicity became unacceptable (>30% weight loss). The reverse schedule, in which CDDP preceded dFdC, did not lead to an increased antitumour effect or to increased toxicity. Adding amifostine, a selective chemoprotector, to the treatment decreased toxicity of the combination without affecting the antitumour effect. Increasing the CDDP dose to 9 mg/kg (day 0) under amifostine protection led to an improved therapeutic index.     

Presence of suppressor cells in spleens of mice bearing a weakly immunogenic syngeneic tumor.

Spleen cells of C57BL/6J mice bearing a poorly immunogenic syngeneic tumor T241 have been shown to suppress the mitogen-induced proliferative responses of normal spleen cells. However, no suppressive effect of these cells was observed on the generation of cytotoxic cells following immunization in vitro against H-2 histocompatibility antigens. The suppressor activity disappeared rapidly after the removal of the primary tumor. Spleen cells of tumor-bearing mice also suppressed the mitogen-induced stimulation of normal spleen cells of mice of different H-2 loci. Removal of phagocytic cells with carbonyl iron treatment had very little effect on the suppressor activity. Suppressor activity was enhanced following fractionation of cells through nylon wool columns. The suppressor population was found to resist anti-immunoglobulin serum and complement treatment, but treatment with anti-thymocyte serum and complement drastically reduced the suppressor activity. These results indicate that cells with suppressor activity have characteristics of T-lymphocytes.     

Increase in alkaline phosphatase activity in the liver of mice bearing Ehrlich ascites tumor.

In mice bearing Ehrilich ascites tumors, alkaline phosphatase activity was increased fivefold in the liver and by 50% in the kidney. In mice bearing solid tumors caused by inoculation of tumor cells into the axillary region, the activity of this enzyme in the liver was increased 11-fold, whereas the activity in the kidney did not change. Alkaline phosphatase activities in the liver and kidney were not altered by administration of adrenal steroids. Adrenalectomy, fasting, and pregnancy did not affect the activity of alkaline phosphatase in the liver and kidney. Treatment with tumor extracts or ascites fluid of normal mice increased liver alkaline phosphatase activity. These findings suggested that the elevation of liver alkaline phosphatase activity was cuased primarily by the tumor itself, and not by hormonal imbalance provoked secondarily by the presence of the tumor.     

Protein synthesis in liver and skeletal muscle of mice bearing an ascites tumor

In mice bearing an ascites tumor at an advanced stage of growth, the weight of the gastrocnemius muscle fell, whereas that of the liver increased. Fractional rates of protein synthesis were measured in vivo under conditions designed to minimize uncertainties in the determination of the specific radioactivity of the precursor amino acid pool. Protein synthesis in liver increased in the tumor-bearing mice in comparison with controls either fed ad libitum or pair-fed to the reduced food intake of the tumor-bearing group. In muscle, the rate of protein synthesis fell substantially in comparison to ad libitum-fed controls but was not significantly different from that in a group for which food intake was restricted to that of the tumor-bearing animals.     

Lipogenesis in tumour and host tissues in mice bearing colonic adenocarcinomas.

Although animals bearing the MAC16 colon adenocarcinoma showed progressive weight loss, the average food consumption (15.1 +/- 0.6 Kcal day-1) did not differ from non tumour-bearing controls (15.3 +/- 0.3 Kcal day-1), while animals bearing a related colon adenocarcinoma, MAC13, which had no effect on body weight had a significantly (P less than 0.01) elevated food intake (16.4 +/- 0.3 Kcal day-1) above controls. Weight loss in animals bearing the MAC16 tumour was associated with a significant reduction in the percentage contribution of the kidneys, colon and epididymal fat pads to the total body weight. Although loss of body fat occurred only in the MAC16 model, both tumours were capable of synthesising lipids from glucose both in vitro and in vivo at the same rate. In addition both tumours increased the rate of lipogenesis from glucose in kidney, liver and epididymal fat pads of the host. Lipogenesis from glucose would be expected to result in a loss of utilisable carbohydrate energy and thus would be expected to increase the overall energy requirements in the tumour-bearing state leading to catabolism of host body tissues if the energy intake is not increased.     

In vivo production of tumor necrosis factor for the treatment of mice bearing Ehrlich ascites tumor.

Tumor necrosis factor (TNF) was produced in mice bearing Ehrlich ascites tumor (EAT) by priming with zymosan and subsequently challenging with lipopolysaccharide. The optimal conditions for the in vivo production of TNF in treating EAT bearing mice were established. The endotoxin shock induced in mice during TNF production could be minimized by the combined administration of sulindac and mannoheptulose. The endogenous TNF produced could suppress proliferation of EAT cells as well as prolong the survival time of mice bearing small tumors.