Norimasa Shudo

Novel screening method for agents that overcome classical multidrug resistance in a human cell line

P-glycoprotein (P-gp) is involved in the transport of a wide variety of organic compounds including a fluorescent dye, rhodamine 6G (RG), as well as anti-cancer drugs. Agents that overcome classical multidrug-resistance (MDR) increased the accumulation of RG in a MDR cell line, KB-C1. The effect of agents on RG-accumulation in KB-C1 cells was highly correlated with their effect on the reversion of vincristine-resistance in KB-C1 cells. RG was detected on a fluorescence microplate reader with a rhodamine channel. This assay may become a useful method for the screening of agents that overcome classical MBR, since it is quick and simple.

Preferential induction of Th17 cells in vitro and in vivo by Fucogalactan from Ganoderma lucidum (Reishi)

The mushroom known as Reishi (Ganoderma lucidum) has been used as an herbal medicine for tumor treatment and immune system activation. Because its effects on the differentiation of effector T helper cells have not yet been fully understood, we investigated the effects of Reishi and those of its principal ingredient, β-glucan, on the activation of dendritic cells and the differentiation of Th17 cells. Reishi extracts as well as purified β-glucan (Curdran) activated DCs and caused them to produce large amounts of IL-23. β-glucan also enhanced and sustained the transcription of IL-23p19. The MEK-ERK signaling pathway positively regulates IL-23p19 transcription in β-glucan-stimulated DCs. In a mixed leukocyte reaction, Reishi-stimulated DCs preferentially induced Th17 cells. Furthermore, orally-administrated Reishi increased the percentages of Th17 cells and the transcription levels of antimicrobial peptides. Our results show that Reishi and β-glucan activate DCs to produce large amounts of IL-23, which induces Th17 differentiation both in vitro and in vivo.

Potentiation of the Vincristine Effect on P388 Mouse Leukemia Cells by a Newly Synthesized Dihydropyridine Analogue, PAK‐200

A newly synthesized dihydropyridine analogue, 2‐[benzyl(phenyl)amino]ethyl 1,4‐dihydrb‐2,6‐dimethyl‐5‐(5,5‐dimethyl‐2‐oxo‐l,3,2‐dioxaphosphorinan‐2‐yl)‐l‐(2‐morpholinoethyl)‐4‐(3‐nitrophen‐yl)‐3‐pyridinecarboxylate (PAK‐200), at 1 μM completely reversed the resistance to vincristine in vincristine‐resistant P388 mouse leukemia cells (P388/VCR), in vitro. PAK‐200 at 2 μM inhibited the efflux of [3H]vincristine from P388/VCR and increased the accumulation of [3H]vincristine in P388/VCR to a level similar to that in P388 cells. P‐Glycoprotein in membrane vesicles from P388/ VCR cells was photolabeled with [3H]azidopine. The labeling was completely inhibited by 10 μM PAK‐200. The calcium antagonistic activity of PAK‐200 was about 1000 times lower than that of another dihydropyridine analogue, nicardipine. Experiments with P388 and P388/VCR‐bearing mice showed that PAK‐200 enhanced the effect of vincristine on both leukemia cells in vivo. These results suggest that PAK‐200 interacts with P‐glycoprotein and reverses drug resistance in P388 mouse leukemia cells in vitro, and that PAK‐200 has an ability to potentiate the effect of vincristine on P388 mouse leukemia cells in vivo.