Effect of a synthetic lipid immunomodulator on the regulation of the transcription factor NF-kappaB.

Macrophage activation plays a central role in host defense against a variety of pathogens via inducible messengers. The transcription factor NF-kappaB controls the synthesis of cytokines involved in immune responses. In quiescent cells, NF-kappaB is located in the cytosol bound to an inhibitor IkappaB. Upon appropriate signal, NF-KB translocates to the nucleus and binds to DNA. The present study investigated the involvement of an immunomodulator, (diHDA-glycerol) on the NF-kappaB/IkappaB complex. Results were compared to those obtained with lipopolysaccharide (LPS) as a major virulence factor in bacterial sepsis. Data showed that exposure of J774.1 cells either to LPS or diHDA-glycerol substantially increased with time the nuclear levels of NF-kappaB complexes. Antibodies to various NF-kappaB proteins supershifted p50, p65 and to a lesser extent c-rel. Western blot analyses showed a rapid cytosolic IkappaB-alpha turn over following LPS exposure in contrast to diHDA-glycerol treatment. Further experiments investigated the involvement of protein kinase C (PKC) by using two inhibitors, staurosporine and H7. Pretreatment of J774.1 with either inhibitor prior to diHDA-glycerol or LPS exposure decreased NF-kappaB activation. Our results indicate that diHDA-glycerol was acting on NF-kappaB through IkappaB regulative mechanisms differing from those used by LPS. DiHDA-glycerol is likely acting on many other transcription factors targeting distinct genes implied in up regulation of the immune system.

In vitro biological activities of arglabin, a sesquiterpene lactone from the Chinese herb Artemisia myriantha Wall. (Asteraceae).

The immunomodulating properties of arglabin, a sesquiterpene lactone isolated from Artemisia myriantha Wall. (Asteraceae) were investigated using the murine macrophage tumor line J774.1. Arglabin-stimulated macrophages displayed a strong cytotoxic activity and the lowest doses (1.25 micrograms/mL and 0.125 micrograms/mL) induced a significant stimulation of cell mitochondrial metabolism, which correlated with [3H]TdR uptake by J774.1 cells under the same experimental conditions. In addition, the secretion of cytokines involved in host defence mechanisms--IL-1, TNF-alpha, and IL-2--was investigated upon incubation of J774-1 cells with arglabin. Arglabin triggered the production of the three cytokines from J774-1 cells. However, the pattern of cytokine secretion differed to some extent, according to the methodology used for cytokine measurement: either traditional bioassay or specific immunoassay (ELISA). Our data emphasize a possible proliferative effect of arglabin in the traditional bioassays, at least for the highest concentrations used. The results were verified with specific ELISA immunoassays. Using either method, lower concentrations of arglabin (ranging from 12.5 micrograms/mL to 0.125 micrograms/mL) were the most effective in inducing IL-1, TNF-alpha, or IL-2 secretion. In addition, preliminary data on phagocytosis showed that arglabin enhanced the uptake of fluorescent latex beads by J774.1 cells.

Effects of taxol on the macrophage function. Interactions with some immunological parameters.

The effects of Taxol on some immunological parameters were investigated, in vitro, in the murine macrophage cell line J774.1. Mitochondrial dehydrogenase activity and (3H) TdR incorporation were shown to be increased in Taxol treated cells. Likewise, Taxol induced TNF alpha secretion. But Taxol seemed to be a weak inducer of the two interleukins IL-1 and IL-2, since their detection occurred late in the cell culture supernatants.

In vitro and in vivo antitumoral activity of free, and encapsulated taxol.

The anti-tumoral activity of taxol encapsulated either in liposomes or in nanocapsules was compared with that of free taxol, using the P388 and L1210 leukaemia test systems. The in vitro inhibition of cell growth was measured after 48 h and 96 h exposure to various concentrations of taxol. With P388 cells, the inhibitory activities of the three forms of the drug were similar. With the L1210 cells, however, the concentrations required for a 50 per cent inhibition of cell growth (IC50) after 48 h exposure to the drug were greater for nanocapsules than for liposomes or free taxol, the values being 0.060, 0.043 and 0.035 micrograms ml-1, respectively. However, a greater efficiency of nanocapsules was observed after 96 h exposure. Using cytomorphometric analysis, no difference was found between L1210 cells treated either with free or encapsulated taxol. In vivo, mice bearing P388 leukaemia, and treated either with taxol solubilized with 5 per cent DMSO + 5 per cent cremophor in saline solution, or with taxol encapsulated in liposomes (IP daily dose of 12.5 mg Kg-1 body weight x 4 days) showed ILS values of 65.8% and 67.9% respectively. Nanocapsules proved to be toxic, apparently due to their composition: this problem is currently under investigation.

Antineoplastic activity of two taxol derivatives on an ovarian tumor xenografted into nude mice.

In order to compare the antineoplastic activities of taxol A, taxol B, a mixture of the two (taxol A 72%) and vinblastine, a human ovarian tumor serially transplanted into 104 female athymic mice was used. In the first experiment (11th passage), the antineoplastic activities of taxol A, taxol B and the mixture taxol AB were tested. The same dose was used in each case (12.5 mg/kg i.e. 1/20 of the evaluated LD50 value). It was administered subcutaneously for 5 consecutive days. Three courses of treatment were performed, with 2 rest periods of 1 week in between. All the taxol derivatives produced a statistically significant delay in the tumor growth. However, taxol B had the lowest chemotherapeutic response. In the second experiment (18th passage), different dose levels were administered (mixture 12.5 mg/kg/day x 4 - taxol A 8.8. mg/kg/day x 4 - taxol B 3.5 mg/kg/day x 4 - vinblastine 0.5 mg/kg/day x 2). For all the taxol derivatives 4 treatment courses with 3 rest periods of 4 days were used, and for vinblastine 4 treatment courses with 3 rest periods of 1 week. At the end of the second experiment, vinblastine, taxol A and a mixture of the two showed similar significant activity, whereas no objective antitumor response was observed following the taxol B treatment at the dose level chosen. The experimental results obtained clearly demonstrate that, in the taxane system, the greatest degree of antineoplastic activity can be attributed to taxol A.

Therapeutic response to taxol of six human tumors xenografted into nude mice.

To test the antineoplastic activity of taxol, a natural product isolated from yew (Taxus baccata L.), six human tumors transplanted into athymic mice were used (primary tumors of breast, endometrium, ovary, brain, lung and a recurrence of tongue tumor). While the growth rates varied with the histopathological characteristics of different tumor types, all mice were treated at a mean tumor volume of 200 +/- 8 mm3. Taxol was given SC at a dose level of 12.5 mg/kg per injection per day for 5 consecutive days out of 7 over a period of 3 weeks. With this schedule antitumor responses were obtained in all of the six neoplasms xenografted into nude mice. In the case of the ductal carcinoma of the breast total tumor regressions were observed in four of the five treated animals. In the five other experimental models taxol produced significant growth delays. We believe that the results of these initial tests on the nude mouse--human tumor xenograft system are convincing and justify clinical assessment of this drug.

[Action of taxol on human tumors transplanted in athymic mice]. / Action du taxol vis-à-vis de tumeurs humaines transplantées sur des Souris athymiques.

In order to study the antitumour activity of taxol (a diterpene of the taxane type) isolated from Yew: Taxus baccata L. (Taxacae), human tumours implanted as xenografts in athymic Mice were used. Swiss nude mice were treated subcutaneously (12.5 mg/kg/injection/day for 5 consecutive days out of 7, over a period of 3 weeks). Treatment by taxol of a liver metastasis of a breast tumour, a tongue primary tumour and a skin metastasis of bronchial carcinoma gave statistically significant results (0,01 greater than P greater than 0,001 and P less than 0,001). However, taxol showed a very moderate antitumour activity against a transplanted primary tumour of the colon.