Nerve growth factor effects on human and mouse melanoma cell invasion and heparanase production

The role of growth factor networks in regulating the progression of human melanocytes towards tumorigenicity and ultimately the malignant phenotype is poorly understood. In particular, the autocrine and paracrine influences that modulate cellular invasion and extracellular matrix degradative enzymes of melanoma cells remain undefined at the molecular level. We report here that nerve growth factor (NGF) can modify some metastasis-associated cellular properties of human and mouse melanoma cells. Treatment of early-passage human metastatic melanoma cells (MeWo) or their variants (3S5, 70W) with biologically active 2.5S NGF resulted in (a) delayed density-dependent inhibition of melanoma cell growth; (b) increased in vitro invasion through a reconstituted basement membrane; and (c) time- and dose-dependent induction of heparanase, a heparan-sulfate-specific endo-β-D-glucuronidase associated with human melanoma metastasis. These effects of NGF were most marked in the 70W brain-colonizing cells (70W > MeWo > 3S5). The NGF enhancement of heparanase secretion was not species-specific, since it was also observed in murine B16 melanoma cells; the highest NGF stimulation of heparanase was found in brain-colonizing murine B16-B15b variant (B16-B15b > B16-BL6, B16-F10, B16-F1). NGF also increased the invasive capacity of the human 70W and murine B16-B15b sublines in a chemoinvasion assay performed with filters coated with purified heparan sulfate proteoglycan (HSPG). The enhancement of chemotactic response and heparanase production was detected at NGF concentrations sufficient to fully saturate both low- and high-affinity NGF receptors (NGFR), the neurotrophin receptor (p75) and the trkA gene product, respectively. The results suggest that, in addition to the effects of NGF on cellular development and differentiation within the peripheral and central nervous systems, NGF can exert changes in the invasive properties of neuroectoderm-derived melanoma cells.     

Randomized trial of paclitaxel plus supportive care versus supportive care for patients with advanced non-small-cell lung cancer

BACKGROUND: In phase II trials, paclitaxel has been shown to have antitumor activity in patients with advanced non-small-cell lung cancer (NSCLC). However, the survival and quality-of-life (QOL) benefits of paclitaxel used as a single agent compared with supportive care alone have not been assessed in a randomized clinical trial. METHODS: A total of 157 patients with stage IIIB or IV NSCLC who had received no prior chemotherapy were randomly assigned to receive either best supportive care alone (78 patients) or paclitaxel plus supportive care (79 patients). Paclitaxel was administered as a 3-hour intravenous infusion every 3 weeks. Supportive care included palliative radiotherapy and supportive therapy with corticosteroids, antibiotics, analgesics, antiemetics, transfusions, and other symptomatic therapy as required. The primary end point of the study was survival. Time to disease progression, response rate, adverse events, and QOL were secondary end points. RESULTS: Pretreatment characteristics were evenly distributed between the two arms. Survival was statistically significantly better in the paclitaxel plus supportive care arm than in the supportive care alone arm (two-sided P =.037) (median survival = 6.8 months versus 4.8 months). Cox multivariate analysis showed paclitaxel plus supportive care to be statistically significantly associated with improved survival (two-sided P =.048). QOL was similar for both treatment arms, except for the functional activity score of the Rotterdam Symptom Checklist, where QOL data statistically significantly favored the paclitaxel plus supportive care arm (two-sided P =.043). CONCLUSION: The addition of paclitaxel to best supportive care significantly improved survival and time to disease progression compared with best supportive care in patients with advanced NSCLC and may improve some aspects of QOL.     

Differential stimulation of the growth of lung-metastasizing tumor cells by lung (paracrine) growth factors: identification of transferrin-like mitogens in lung tissue-conditioned medium.

Certain metastatic tumor cells successfully form metastases at particular organ sites, and their organ colonization properties cannot be explained by mechanical or anatomic factors. These tumor cells possess the ability to colonize such sites through preferential adhesion to organ microvessel endothelial cells, preferential organ invasion by expression of particular degradative enzymes and response to organ motility factors, and preferential organ growth by response to growth factors present at relatively higher concentrations in the target organ. The likelihood that target organ-associated growth factors exist and are important in metastatic colonization has been approached by studying the mitogenic effects of target organ extracts, fragments, or conditioned media on poorly and highly metastatic tumor cells that show organ preference of metastasis. We previously described the isolation of a major organ-derived (paracrine) growth factor from lung tissue-conditioned medium. Characterization of this mitogen has demonstrated that it is a transferrin or a transferrin-like glycoprotein, and antibodies to transferrin can remove significant growth activity from lung tissue-conditioned medium. Further demonstration of the existence and characterization of metastasis-associated organ (paracrine) growth factors and their receptors will be helpful in understanding the organ preference of metastasis.     

Tumor-cell-platelet aggregation does not correlate with metastatic potential of rat 13762NF mammary adenocarcinoma tumor cell clones

The ability of tumor cells to induce platelet aggregation has been correlated with their capacities to colonize the lungs of experimental animals. We tested this hypothesis by studying the ability of cloned, low-passage metastatic tumor cell lines derived from rat 13762NF mammary adenocarcinoma to aggregate rat platelets in vitro and in vivo, and we then compared this activity to metastatic potential by determining the incidence of lung metastasis after subcutaneous or intravenous inoculation of the tumor cell clones into syngeneic rats. Our results failed to show a correlation between in vitro platelet-aggregating activity and metastatic potential. In this system we could not detect platelet-aggregation activity with the most metastatic tumor clone, while the least metastatic clone clearly possessed high platelet-aggregating activity. In addition, by measuring changes in blood platelet and fibrinogen concentrations at various intervals following intravenous injection of tumor cell clones, we were able to confirm in vivo the observed in vitro differences in their plateletaggregating activities. Thus, platelet aggregating activity is heterogeneously expressed among 13762NF cell clones and appears unrelated to spontaneous or experimental metastasis in this tumor system.     

Boomslang bite with haemorrhage and activation of complement by the alternate pathway

A case of Boomslang poisoning is reported for the first time in North West Europe. The treatment is described and activation of complement by the alternate pathway in vivo was shown.     

Lipid replacement/antioxidant therapy as an adjunct supplement to reduce the adverse effects of cancer therapy and restore mitochondrial function

The most common complaints of cancer patients undergoing chemoor radiotherapy are fatigue, nausea, vomiting, malaise, diarrhea and headaches. These adverse effects are thought to be due to damage of normal tissues during the course of therapy. In addition, recent evidence indicates that fatigue is related to reduced mitochondrial function through loss of efficiency in the electron transport chain caused by membrane oxidation, and this occurs during aging, in fatiguing illnesses and in cancer patients during cytotoxic therapy. Lipid Replacement Therapy administered as a nutritional supplement with antioxidants can prevent oxidative membrane damage to normal tissues, restore mitochondrial and other cellular membrane functions and reduce the adverse effects of cancer therapy. Recent clinical trials using patients with chronic fatigue have shown the benefit of Lipid Replacement Therapy plus antioxidants in restoring mitochondrial electron transport function and reducing moderate to severe chronic fatigue by protecting mitochondrial and other cellular membranes from oxidative and other damage. In cancer patients a placebo-controlled, cross-over clinical trial using Lipid Replacement Therapy plus antioxidants demonstrated that the adverse effects of chemotherapy can be reduced in 57–70% of patients. Dietary use of unoxidized membrane lipids plus antioxidants is recommended for patients undergoing cancer therapy to improve quality of life but should not be taken at the same time of day as the therapy. The author has no financial interest in the products discussed in this contribution.     

Report of two protocol planned interim analyses in a randomised multicentre phase III study comparing capecitabine with fluorouracil and oxaliplatin with cisplatin in patients with advanced oesophagogastric cancer receiving ECF

The purpose of the study was to establish the optimal dose of capecitabine (X) to be used within a multicentre, randomised study evaluating the potential roles of oxaliplatin (O) and X in chemonaive patients (pts) with advanced oesophagogastric cancer. Two by two design was used, and pts were randomised to one of four regimens and stratified for extent of disease, performance status (PS) and centre. The treatment regimens are epirubicin, cisplatin, 5-fluorouracil (ECF), EOF, ECX or EOX. Doses: E 50 mg m(-2), C 60 mg m(-2) and O 130 mg m(-2) i.v. 3 weekly; F 200 mg m(-2) day(-1) i.v. and X 500 mg m(-2) b.i.d.(-1) (escalated to 625 mg m(-2) b.i.d.(-1) after results of first interim analysis) p.o., continuously. First interim analysis was performed when 80 pts had been randomised. Dose-limiting fluoropyrimidine toxicities were stomatitis, palmar plantar erythema (PPE) and diarrhoea; 5.1% of X-treated pts experienced grade 3/4 toxicity. Protocol planned dose escalation of X to 625 mg m(-2) b.i.d.(-1) was instituted and a second interim analysis has been performed; results are presented in this paper. A total of 204 pts were randomised at the time of the protocol planned 2nd interim analysis. Grade 3/4 fluoropyrimidine-related toxicity was seen in 13.7% pts receiving F, 8.4% pts receiving X 500 mg m(-2) b.i.d.(-1) and 14.7% pts receiving X 625 mg m(-2) b.i.d.(-1). Combined complete and partial response rates were ECF 31% (95% CI 18.7-46.3), EOF 39% (95% CI 25.9-53.1), ECX 35% (95% CI 21.4-50.3), EOX 48% (95% CI 33.3-62.8). Grade 3/4 fluoropyrimidine toxicity affected 14.7% of pts treated with X 625 mg m(-2) b.i.d.(-1), which is similar to that observed with F, confirming this to be the optimal dose. The replacement of C by O and F by X does not appear to impair efficacy. The trial continues to total accrual of 1000 pts.     

Generation of influenza vaccine viruses on Vero cells by reverse genetics: an H5N1 candidate vaccine strain produced under a quality system

Human influenza vaccine reference strains are prepared as required when an antigenically new strain is recommended by WHO for inclusion in the vaccine. Currently, for influenza A, these strains are produced by a double infection of embryonated hens' eggs using the recommended strain and the laboratory strain PR8 which grows to high titre in eggs, in order to produce a high growth reassortant (HGR). HGRs are provided by WHO reference laboratories to the vaccine manufacturing industry which use them to prepare seed virus for vaccine production. The use of reverse genetics in preparing vaccine reference strains offers several advantages over the traditional method: (i) the reverse genetics approach is a direct rational approach compared with the potentially hit-or-miss traditional approach; (ii) reverse genetics will decontaminate a wild type virus that may have been derived in a non-validated system, e.g. a cell line not validated for vaccine purposes, or that may contain additional pathogens; (iii) at the plasmid stage, the HA can be engineered to remove pathogenic traits. The use of reverse genetics in deriving HGRs has been demonstrated by several laboratories, including its use in deriving a non-pathogenic reassortant strain from a highly pathogenic virus. In this report, we have advanced the use of reverse genetics by making use of a cell line acceptable for human vaccine production, by demonstrating directly the short time frame in which a reassortant virus can be derived, and by deriving a non-pathogenic pandemic vaccine reference virus in cells validated for vaccine production and under quality controlled conditions.     

A vector expressing feline mature IL-18 fused to IL-1beta antagonist protein signal sequence is an effective adjuvant to a DNA vaccine for feline leukaemia virus

DNA vaccination using vectors expressing the gag/pol and env genes of feline leukaemia virus (FeLV) and plasmids encoding feline interleukin-12 (IL-12) and IL-18 completely protected cats from viraemia following challenge [Hanlon L, Argyle D, Bain D, Nicolson L, Dunham S, Golder MC, et al. Feline leukaemia virus DNA vaccine efficacy is enhanced by coadministration with interleukin-12 (IL-12) and IL-18 expression vectors. J Virol 2001;75:8424-33]. However, the relative contribution of each cytokine gene towards protection is unknown. This study aimed to resolve this issue. IL-12 and IL-18 constructs were modified to ensure effective expression, and bioactivity was demonstrated using specific assays. Kittens were immunised intramuscularly with FeLV DNA and various cytokine constructs. Together with control kittens, these were challenged oronasally with FeLV and monitored for 15 weeks. All six kittens given FeLV, IL-12 and IL-18 were protected from the establishment of persistent viraemia and four from latent infection. Of six kittens immunised with FeLV DNA and IL-18, all were protected from viraemia and five from latent infection. In contrast, three of five kittens given FeLV DNA and IL-12 became persistently viraemic. Therefore, the adjuvant effect on the FeLV DNA vaccine appears to reside in the expression of IL-18.