Alterations in proteolytic activity at low pH and its association with invasion: a theoretical model.

The extracellular pH (pHe) of solid tumours is often lower than in normal tissues, with median pH values of about 7.0 in tumours and 7.5 in normal tissue. Despite this more acidic tumour microenvironment, non-invasive measurements of intracellular pH (pHi) have shown that the pHi of solid tumours is neutral or slightly alkaline compared to normal tissue (pHi 7.0-7.4). This gives rise to a reversed cellular pH gradient between tumours and normal tissue, which has been implicated in many aspects of tumour progression. One such area is tumour invasion: the incubation of tumour cells at low pH has been shown to induce more aggressive invasive behaviour in vitro. In this paper the authors use mathematical models to investigate whether altered proteolytic activity at low pH is responsible for the stimulation of a more metastatic phenotype. The authors examined the effect of culture pH on the secretion and activity of two different classes of proteinases: the metalloproteinases (MMPs), and the cysteine proteinases (such as cathepsin B). The modelling suggests that changes in MMP activity at low pH do not have significant effects on invasive behaviour. However, the model predicts that the levels of active-cathepsin B are significantly altered by acidic pH. This result suggests a critical role for the cysteine proteinases in tumour progression.

Stereo-specific cytotoxic effects of gossypol enantiomers and gossypolone in tumour cell lines.

The naturally occurring compound, gossypol, has been previously used as a male oral contraceptive, for the treatment of benign gynaecological conditions and cancer patients. Long-term daily dosing with gossypol is associated with minimal side effects and no myelosuppression. Since gossypol exhibits atropisomerism due to the restricted rotation about the 2,2' carbon bond, we have isolated the l- and d-isomers by Schiff's base formation using a chiral amine and regenerated the enantiomers by acid hydrolysis. The enantiomers and the proposed oxidative metabolite, gossypolone, were characterized by HPLC, 1H-NMR and optical rotation. The cytotoxicity was assessed in cell cultures derived from melanoma, lung, breast, cervix, and leukaemia using the MTT viability assay. The cytotoxicity of gossypolone was similar to racemic gossypol in five out of the six cell lines studied. The l-enantiomer of gossypol induced a dose-dependent cell kill in all cell lines with a mean IC50 of 20 microM and was significantly more potent than racemic gossypol, the d-enantiomer of gossypol and gossypolone. In addition, when the leukaemia line was exposed to l-gossypol (0.5-10 microM) over a 4-day period, a schedule-dependent decrease in cell viability was observed. l-Gossypol was also compared with respective drugs used to treat patients with melanoma, lung cancer and leukaemia. The data indicate that l-gossypol was significantly more active than cisplatin, melphalan and dacarbazine in the two melanoma lines, cisplatin and daunorubicin in the lung line and hydroxyurea and busulphan in the leukaemia line. Preliminary studies using one melanoma line showed that the l-isomer induced cell shrinkage, membrane blebbing and DNA fragmentation, characteristics suggestive of apoptotic cell death.

Modelling tumour acidity and invasion.

The intracellular pH (pHi) of mammalian cells is tightly regulated by the concerted action of a number of different pumps in the plasma membrane. Despite the acidic extracellular environment (pHe 6.8-7.0) of some tumours, the pHi of solid tumours is neutral or slightly alkaline compared to normal tissues (pHi 7.0-7.4). This gives rise to a reversed pH gradient across the cell membrane between tumours and normal tissue, which has been implicated in many aspects of tumour progression. One such area is tumour invasion: the incubation of tumour cells at low pH has been shown to induce more aggressive invasive behaviour in vitro. In this paper we use mathematical models to investigate whether altered proteolytic activity at low pH is responsible for the stimulation of a more metastatic phenotype. We examined the effect of culture pH on the secretion and activity of two different classes of proteinases: the metalloproteinases (MMPs), and the cysteine proteinases (such as cathepsin B). The modelling predicts that, in addition to metalloproteinase activity, the pH-induced peripheral redistribution of cathepsin B could be a major factor involved in the acquisition of a more metastatic phenotype in malignant cells at low pHe.

Changes in serum acute phase proteins in breast cancer patients receiving methotrexate infusion therapy.

The serum concentrations of six glycoproteins (alpha 1-acid glycoprotein, AGP; haptoglobin, Hp; alpha 1-antitrypsin, AT; ceruloplasmin, Cp; prealbumin, PALB; and alpha 2-macroglobulin, MACRO) have been estimated serially in nine advanced breast cancer patients who received a total of 24 intravenous infusions of methotrexate (MTX). The serum glycoprotein levels taken before the first drug exposure did not relate with the prognosis of these patients. In eight patients, constantly high or rising levels of AGP, Hp and AT during consecutive infusions of the drug were associated with continued metastatic disease. A transient tumour regression occurred in one patient which correlated with falling serum levels of these proteins into the normal range. The serum levels of Cp, PALB and MACRO did not correlate with the clinical status of these patients during treatment. Possible factors which may influence the serum levels of these glycoproteins in cancer patients during therapy are discussed.

Acute phase glycoproteins in sera of patients with sarcomas receiving methotrexate infusion therapy.

The serum concentrations of alpha-1-acid glycoprotein, haptoglobin and alpha-1-antitrypsin have been estimated serially in eight patients with malignant tumours of bone and soft tissue, who received a total of thirty-four intravenous infusions of high-dose methotrexate. The serum glycoprotein levels taken before the first drug exposure did not relate with the prognosis of these patients. A tumour mass (40 mm) did not influence the serum levels of these glycoproteins. The presence of a pleural effusion was associated with increased serum levels of these glycoproteins. These assays appear to be of no value in monitoring the course of the disease in this group of cancer patients.

Changes in serum acute phase proteins in ovarian cancer patients receiving cis-diamminedichloroplatinum (CDDP) infusion therapy.

Nine patients with adenocarcinoma of the ovary were given a number of courses of Cisplatin, by I.V. infusion. In four patients a complete clinical response was observed and these patients are disease-free one year after treatment. There was no clinical response in the remaining five patients who have subsequently died. Serial determinations of three acute-phase reactant proteins (alpha 1-acid glycoprotein, haptoglobin, alpha 1-antitrypsin) were performed before every infusion and after therapy. Constantly high or rising serum levels of haptoglobin and alpha 1-acid glycoprotein were associated with progression of the cancer, whereas in patients who were disease-free after therapy, these glycoproteins remained essentially in the normal range. The results suggest that serial measurements of haptoglobin and alpha 1-acid glycoprotein may have clinical value as aids in deciding the effectiveness of drug therapy in these patients.

Cytotoxicity of gossypol enantiomers and its quinone metabolite gossypolone in melanoma cell lines.

The cytotoxicity of the (-)- and (+)-isomers and the quinone metabolite gossypolone prepared from the naturally occurring polyphenolic dialdehyde gossypol were compared using two human melanoma cell lines (SK-mel-19 and SK-mel-28) with a similar growth rate, one melanotic (melanin content of 69 pg/cell) and one amelanotic (melanin content of 10 pg/cell). Results from two viability assays (MTT and flow cytometry) showed that the cytotoxicity of racemic gossypol was identical for both cell lines (50% inhibition of cell growth IC50 = 22 microM). Gossypolone at equimolar concentrations was inactive in the amelanotic cell line and as potent as racemic gossypol in the melanotic cell line. (-)-Gossypol was significantly more active in both cell lines compared with the (+)-isomer. The cytotoxic effect of (-)-gossypol was both concentration and time dependent. Under serum-free conditions, the cytotoxicity of both enantiomers was increased, suggesting that serum protein binding may play a role in the differential toxicity of these isomers in vitro. Morphological changes after exposure to (-)-gossypol included shrinkage and loss of adherence. Cell sensitivity to the (-)-isomer was five-fold greater (IC50 = 4 microM) using a clonogenic assay. At equimolar concentrations, (-)-gossypol was more cytotoxic to both cell lines than the clinically used drugs cisplatin, dacarbazine and melphalan. The results of this study suggest that (-)-gossypol may be of potential therapeutic benefit in melanoma patients.

Evaluation of an immunoradiometric assay for the detection of an ovarian tumour marker, CA125, in serum.

An immunoradiometric assay, that uses a monoclonal antibody attached to a solid support and developed for the detection of the ovarian tumour antigen, CA125, in serum, has been evaluated. The method was simple to perform, with reagents having a shelf-life of approximately 4 weeks. The potentials and limitations of this test are discussed.

Intravesical Bacillus Calmette-Guerin in Ta and T1 Bladder Cancer.

BACKGROUND: Intravesical therapy with Bacillus Calmette-Guerin (BCG) aims to reduce the incidence of tumour recurrence following transurethral resection (TUR) for patients with superficial bladder cancer. OBJECTIVES: The objective of this review was to compare the incidence of tumour recurrence after the standard therapy of transurethral resection versus transurethral resection plus intravesical Bacillus Calmette-Guerin. SEARCH STRATEGY: We searched the Cochrane Controlled Trials Register (March 2000), Medline (February, 2000), EMBASE (February, 2000), Cancerlit (February, 2000), Healthstar (February, 2000), Database of Abstracts of Reviews of Effectiveness (February, 2000) and the Bath Information Data Service. The Proceedings of the American Society Clinical Oncology was hand searched (1996 - 1999). SELECTION CRITERIA: Randomised or quasi-randomised trials of transurethral resection alone versus transurethral resection plus intravesical Bacillus Calmette-Guerin. Patients with Ta and T1 bladder cancer of medium or high risk of tumour recurrence, were eligible for inclusion. DATA COLLECTION AND ANALYSIS: Four reviewers assessed trial quality and two abstracted the data independently. The Peto odds ratios and log hazard ratios were determined to compare the number of patients with disease recurrence at 12 months and the rate of recurrence, respectively. MAIN RESULTS: Six randomised trials were included involving 585 eligible patients. There were significantly fewer patients with disease recurrence at 12 months in the BCG plus TUR group compared to those that received TUR alone (odds ratio 0.30, CI 0.21, 0.43). The overall log hazard ratio for recurrence (-0.83, variance 0.02) indicated a significant benefit of BCG treatment in reducing tumour recurrence. Toxicities associated with BCG consisted mainly of cystitis (67%), haematuria (23%), fever (25%) and urinary frequency (71%). No BCG-induced deaths were reported. REVIEWER'S CONCLUSIONS: In patients with medium/high risk Ta or T1 bladder cancer, immunotherapy with intravesical BCG following TUR appears to provide a significant advantage over TUR alone in delaying tumour recurrence.

OC125 immunoscintigraphy in ovarian carcinoma: a comparison with alternative methods of assessment.

The membrane bound, tumour associated antigen CA125 is recognized by the monoclonal antibody OC125 and may be detected in tumour tissue and serum in over 80% of patients with epithelial ovarian carcinomas. A total of 13 immunoscintigrams using 111 MBq 131I-OC125 have been performed in 11 patients. The results have been compared with clinical examination, CT and ultrasound scans, surgical findings and serum CA125 concentrations. Macroscopic disease was present at the time of scanning in 11 patients (less than 2 cm, eight patients, greater than 2 cm, three patients). Clinical examination and ultrasound were positive in three, CT scanning in four, immunoscintography in seven and serum CA125 in eight patients. This pilot study suggest that serum CA125 estimation is the most sensitive indicator of disease activity. However, immunoscintigraphy using this agent may localize residual disease when clinical examination and other radiological investigations fail.

Role of gangliosides in tumour progression: a molecular target for cancer therapy?

In a number of patients with tumours of either neuroectodermal or epithelial origin, polysialylated gangliosides (e.g. GD3) are over-expressed. The mechanism of ganglioside over-expression may be different for the two classes of tumour and could represent distinct secondary genetic mutations or epigenetic changes affecting the enzymes (transferases and/or hydrolases) controlling the metabolic interconversions of these gangliosides. Tumour cells of neuroectodermal origin (e.g. melanomas and brain tumours) are known to produce and shed polysialylated gangliosides, whereas paracrine signal(s) from tumour cells of epithelial origin (e.g. carcinomas of cervix, lung, prostate, breast, head and neck, colon and ovary) may stimulate over-expression and shedding from tumour infiltrating mesenchymal cells (e.g. macrophages and/or fibroblasts). This cellular membrane over-expression and shedding of acidic glycosphingolipids into the interstitial spaces and blood of cancer patients may play a central role in increased tumour cell growth, lack of immune cell recognition and neovascularization and could represent a molecular target for cancer therapy.

Biliary excretion of platinum in a patient treated with cis-dichlorodiammineplatinum (II).

Biliary excretion of platinum was measured in a patient receiving an intravenous infusion of cis-dichlorodiammineplatinum (II). Over a 3-day period, less than 1% of the administered platinum was detected in the bile, and 47% appeared in the urine, indicating that biliary excretion is a minor route of elimination.