Objective Investigated the treatment effect of ISCOM leukemia vaccine combination with 1-methyl tryptophan on tumor burden mice .Methods Saponin was added lipase protein (1 mg/mL) 7 ℃ for 12 h ,adding 80 μL lipid mixed solution and 5 mL saponin solution (1 mg/mL ) to prepare ISCOM leukemia vaccine .C57BL /6 mice were randomly divided into model group , ISCOM leukemia vaccine group ,1-methyl tryptophan group and combination group ,Mice were injected FBL-3 cell to built leukemia tumor-burdened mice model .After treatment for 4 weeks ,tumors weight ,NK and Mφ and CTL cell killing activity ,serum levels of IL-10 ,IL-12 were detected .Results Tumor weight in combination group was less than 1-methyl tryptophan and ISCOM leukemia group [(0 .64 ± 0 .26)g vs .(2 .49 ± 0 .91)g ,P< 0 .01 ,(0 .64 ± 0 .26)g vs .(1 .28 ± 0 .73)g ,P< 0 .05] ;NK cell killing activity in com-bination group was higher than 1-methyl tryptophan group[(38 .41 ± 8 .27)% vs .(67 .22 ± 12 .74)% ,all P< 0 .01)] ;M φ activity in combination group was significantly higher than 1-methyl tryptophan group[(55 .69 ± 13 .69)% vs .(69 .47 ± 14 .79)% ,P< 0 .01] ;CTL activity in combination group was significantly higher than 1-methyl tryptophan group and ISCOM leukemia group[(43 .77 ± 8 .89)% vs .(69 .68 ± 11 .44)% ,P< 0 .01 ,(58 .87 ± 9 .45)% vs .(69 .68 ± 11 .44)% ,P < 0 .05] ;IL-10 in combination group were significantly lower than 1 - methyl tryptophan group and ISCOM leukemia group [(76 .2 ± 6 .82)pg /L vs .(98 .3 ± 13 .4)pg/L ,P<0 .01 ,(76 .2 ± 6 .82)pg/L vs .(202 .3 ± 44 .5)pg/L ,P < 0 .01] ;IL-12 in combination group were significantly higher than 1-methyl tryptophan group and ISCOM leukemia group[(381 .2 ± 47 .3)pg/L vs .(332 .1 ± 30 .2)pg/L ,P < 0 .05 ,(381 .2 ± 47 .3)pg /L vs . (291 .2 ± 17 .3)pg/L ,P< 0 .01] .Conclusion Combination with 1-methyl tryptophan and ISCOM leukemia vaccine has a well anti-tumor effect ,its mechanism may be through mediated and the expression of IL-12 and IL-10 .

OBJECTIVE: To explore the exact mechanisms of programmed cell death (PCD) induced by Type II anti-CD20 mAb in CD20+ non-Hodgkin lymphoma (NHL) cells, and to provide theoretical basis for anti-tumor ability of new CD20 mAb.
 METHODS: After incubation with Rituximab (a Type I anti-CD20 mAb) and Tositumomab (a Type II anti-CD20 mAb), Raji cells were stained by annexin V & propidium iodide (PI). The ratio of programmed death cells were measured by two channel flow cytometry (FCM). Before the treatment of anti-CD20 mAbs, Raji cells was incubated with a caspase inhibitor carbobenzoxy-valyl-alanyl-aspartyl-[O-methyl]- fluoromethylketone (Z-VAD-FMK) and a dihydroceramide synthase inhibitor fumonisin B1 (FB1) for 30 minutes to assess their inhibitory effect on PCD. High performance liquid chromatography (HPLC) was utilized to compare the ratio of programmed death cells between the pretreatment group (treated by Rituximab and Tositumomab) and the non-pretreatment group. The anti-CD20 mAbs-treated Raji cells were collected, and the ceramide levels in the Raji cells in the different pretreatment groups were also examined by HPLC, and the inhibitory effect of FB1 on the changes of ceramide levels in the Raji cells was measured. The Raji cells were incubated with different concentration C2-ceramide, C2-Ceramide-induced PCD was also evaluated by annexin V & PI staining after 16 hours. 
 RESULTS: Tositumomab (10 µg/mL) but not Rituximab (10 µg/mL) can induce significant PCD (28.6±4.2)% in Raji cells, with significant difference (t=26.48, P<0.01), which cannot be blocked by Z-VAD-FMK with a concentration range from 10 to 30 µmol/L (F=3.01, P>0.05). The cellular ceramide levels in Raji cells were significantly elevated after the treatment of Tositumomab (t=28.48, P<0.01). C2-ceramide can significantly induce PCD in Raji cells in a dose-dependent manner with a concentration range from 5 to 40 µmol/L (F=2.71, P>0.05). The dihydroceramide synthase inhibitor FB1 can significantly inhibit the elevated cellular ceramide levels (F=20.18, P<0.01) and cell programmed death induced by Tositumomab (F=17.02, P<0.01).
 CONCLUSION Type II but not Type I anti-CD20 mAbs can induce caspase independent PCD in CD20+ NHL cells through the elevation of cellular ceramide levels. The PCD is not associated with classic caspase pathway.
Amino Acid Chloromethyl Ketones ; Apoptosis ; drug effects ; Cell Line, Tumor ; drug effects ; Humans ; Lymphoma, Non-Hodgkin ; Rituximab ; pharmacology ; Sphingosine ; analogs & derivatives ; pharmacology