Synthesis and antitumor activity of duocarmycin derivatives: modification at the C-7 position of segment-A of A-ring pyrrole compounds

A series of the C7-substituted A-ring pyrrole derivatives of duocarmycin were synthesized, and evaluated for in vitro anticellular activity against HeLa S3 cells and in vivo antitumor activity against murine sarcoma 180 in mice. All of the C7-substituted A-ring pyrrole compounds decreased potency in vitro and in vivo. However, some showed strong antitumor activity with T/C values less than 0.3. Among them, the 7-formyl compound 5d showed remarkable potent in vivo antitumor activity and low peripheral blood toxicity, which were equal to 2c.     

三种药用植物内生菌的分离及其抗肿瘤活性菌株的筛选

对204株分离自长春花、海南粗榧和见血封喉的内生菌进行了抗肿瘤活性筛选。结果显示,有19株内生菌的发酵液至少对一种指示瘤株具有细胞毒活性,其中4株菌的发酵液对S180的抑制率较强,均在70%以上。研究表明菌株PA09006和PA09009发酵液中的抗肿瘤活性物质,具有一定的酸碱耐受性,并且对温度和紫外线具有一定的稳定性。     

Compositions and Anti-Tumor Activity of Pyropolyporus fomentarius Petroleum Ether Fraction In Vitro and In Vivo

The chemical compositions and anti-tumor activities of the petroleum ether fraction (PE), from mushroom Pyropolyporus fomentarius, were studied. Upon gas chromatography–mass spectrometry (GC–MS) analysis, nine major constituents were identified in the fraction. In vitro, the PE showed cytotoxic activity against murine sarcoma S180 (S180) cells in a dose- and time-dependent manner, and the cytotoxic effects were associated with apoptosis. The mitochondrial membrane potential loss and the intracellular ROS generation were greatly increased in the Pyropolyporus fomentarius PE treated group, suggesting cell apoptosis, induced by the PE in S180 cells, might be mitochondria dependent and ROS mediated. Consistent with in vitro findings, the in vivo study showed that the Pyropolyporus fomentarius PE was also effective in inhibiting the tumor growth induced by S180 cells and had lower immune organ toxicity. We found that the Pyropolyporus fomentarius PE has significant anti-tumor activity and great potential in screening anti-tumor drugs.     

Purine metabolic enzymes in lymphocytes. II. Adenosine deaminase and purine nucleoside phosphorylase activities in the immune response

ICR mice were immunized with sheep red blood cells (sRBC). Both adenosine deaminase (ADA) and purine nucleoside phosphorylase (PNP) activities in spleen lymphocytes increased faster than the serum antibody titer and reached a peak one week after the immunization. ADA activity increased significantly in T lymphocytes but not in B lymphocytes collected from the spleens of the immunized mice. A statistically significant increase in PNP activity was found in both T and B lymphocytes from the spleens of the immunized mice. Spleen lymphocytes collected from ICR mice which had been immunized with mitomycin C-treated sarcoma 180 (S180) cells one week earlier showed cytotoxic activity against viable S180 cells. Both ADA and PNP activities in spleen lymphocytes of S180-immunized mice increased significantly, and both activities increased in T lymphocytes prepared from spleen of immunized mice. In contrast, an increase was found in PNP activity but not in ADA activity in B lymphocytes. These results suggest that an increase in both ADA and PNP activities may by necessary for the T-cell response in both humoral and cellular immune responses, and that an increase in PNP activity may be necessary for the B-cell response.     

Immunomodulatory Activity of 3β,6β‐Dihydroxyolean‐12‐en‐27‐oic Acid in Tumor‐Bearing Mice

3β,6β‐Dihydroxyolean‐12‐en‐27‐oic acid ( 1 ) is a pentacyclic triterpenoid isolated from the rhizomes of Astilbe chinensis. To evaluate the in vivo antitumor potential and to elucidate its immunological mechanisms, effect of 1 on the growth of mouse‐transplantable tumors, and the immune response in naive and tumor‐bearing mice were investigated. The mice inoculated with mouse tumor cell lines were orally treated with 1 at the doses of 40, 60, and 80 mg/kg for 10 days. The effects of 1 on the growth of mouse‐transplantable S180 sarcoma and H22 hepatoma, splenocyte proliferation, cytotoxic T lymphocyte (CTL) activity, natural killer (NK) cell activity, and production of interleukin‐2 (IL‐2) from splenocytes in S180‐bearing mice were measured. Furthermore, the effect of 1 on 2,4‐dinitrofluorobenzene (DNFB)‐induced delayed‐type hypersensitivity (DTH) reactions and the sheep red blood cell (SRBC)‐induced antibody response in naive mice were also studied. Compound 1 could not only significantly inhibit the growth of mouse transplantable S180 sarcoma and H22 hepatoma, increase splenocytes proliferation, CTL and NK cell activity, and the level of IL‐2 secreted by splenocytes in tumor‐bearing mice, but also remarkably promote the DTH reaction and enhance anti‐SRBC antibody titers in naive mice. These results suggested that 1 could improve both cellular and humoral immune response, and could act as antitumor agent with immunomodulatory activity.     

小鼠腹腔巨噬细胞被S-O_2—1激活后的抑瘤作用 Ⅰ.小鼠腹腔巨噬细胞的体外细胞毒作用及扫描电镜观察

在六十年代Mathe就用卡介苗来治疗小儿白血病,并获得了良好的效果。以后人们开始重视如何利用微生物佐剂刺激机体的防御机制,增强机体对肿瘤的排斥能力,来达到提高肿瘤治疗效果的目的。     

Antitumor Activity of the Essential Oil from the Leaves of Croton regelianus and Its Component Ascaridole

Croton regelianus Muell. Arg. , popularly known as ‘velame‐de‐cheiro’, is a native plant from the Northeast of Brazil used in folk medicine to treat diseases of different kinds, including malignant tumors. In this study, the in vitro and in vivo antitumor effects of the essential oil from the leaves of C. regelianus and ascaridole, one of the main constituents, were investigated. In vitro, the essential oil and ascaridole displayed cytotoxicity, showing IC₅₀ values in the range of 22.2 to 48.0 μg/ml in HL‐60 and SF‐295 cell lines for the essential oil, and 6.3 to 18.4 μg/ml in HL‐60 and HCT‐8 cells lines for ascaridole, respectively. The in vivo study, using sarcoma 180 as a tumor model, demonstrated inhibition rates of 28.1 and 31.8% for essential oil, at the 50 and 100 mg/kg, while ascaridole inhibition rates were 33.9% at 10 mg/kg and 33.3% at 20‐mg/kg doses. Histopathological examination showed that the organs were only weakly affected by the treatment. In conclusion, ascaridole has an interesting antitumor activity in sarcoma 180 murine model, probably related to the described cytotoxic activity, and, moreover, its presence in the essential oil from the leaves of C. regelianus could explain, at least in part, the ethnopharmacological use of this plant in the treatment of cancer.     

Alpha Sarcin, a New Antitumor Agent: II. Fermentation and Antitumor Spectrum

Aspergillus giganteus MDH 18894 was found to produce alpha sarcin, a new antitumor agent inhibitory to a number of different animal tumors. This culture produced culture filtrates that inhibited mouse sarcoma 180 at dilutions up to 1:32. Optimal fermentation conditions were established for shake flasks and 30-liter and 100-gal (378.5-liter) fermentors. Results from a variety of media were reported; however, the medium which yielded the most reproducible filtrates consisted of 2% corn starch, 1.5% beef extract (Difco), 2% peptone (Difco), and 0.5% sodium chloride. Purified preparations of 800 sarcoma 180 dilution units per mg were inactive against bacteria, yeasts, molds, actinomycetes, and protozoa, but were cytotoxic to several strains of mammalian cells. Alpha sarcin inhibited sarcoma 180 in mice at a dose of 62.5 μg per kg per day. A variety of other animal tumors were also inhibited by this compound.     

Oral administration of Agaricus blazei (H1 strain) inhibited tumor growth in a sarcoma 180 inoculation model.

Agaricus blazei (H1 strain) was tested for its anticancer activity using a sarcoma 180 (S180) inoculation model and the changing patterns of splenocyte subsets were examined. Its hot-water extract was administered orally to ICR and KSN nude mice that were inoculated with S180. The growth of S180 was significantly inhibited in A.blazei treated groups. Pan T cells significantly increased in all treated groups compared to controls, even in KSN nude mice. Splenocyte subset changes were slightly different between ICR and KSN nude mice. This S180 inoculation model proved to be effective in screening the antitumor effect of basidiomycetes and allowed comparisons of immunological cellular changes between the mouse strains.     

蚯蚓提取物对小鼠肿瘤动物模型的研究

目的 :研究蚯蚓提取物 (EFE)的免疫活性及抗肿瘤作用。方法 :采用小鼠移植性肿瘤S1 80 肉瘤及Heps肝癌的动物模型观察其肿瘤抑制作用。结果 :EFE对S1 80 肉瘤和Heps肝癌细胞的抑制率分别为 36 97%和 4 8 55% ;结论 :它对小鼠实体瘤细胞有明显的抑制作用 (P <0 0 1 )并提示对小鼠的细胞和体液免疫功能有显著增强作用。     

Antitumor activity of annonaceous acetogenins in HepS and S180 xenografts bearing mice

Annonaceous acetogenins (ACGs), as one of the most powerful groups of mitochondrial complex I inhibitors, exhibit potent cytotoxic activity against a variety of human tumor cell lines. In this study, the antitumor activities of three main types of ACGs were investigated using S180 and HepS xenografts bearing mice simultaneously. The results revealed that select ACGs suppressed tumor growth in a dose-dependent fashion. Tested ACGs showed more selective antitumor activity against HepS. Furthermore, adjacent bis-THF ACGs were more active than mono-THF and nonadjacent bis-THF ACGs against HepS and S180; nonadjacent bis-THF ACGs were more active than mono-THF ACGs against S180, but mono-THF ACGs were more potent than nonadjacent bis-THF ACGs against HepS.     

Synthesis and antitumor activity of duocarmycin derivatives: A-ring pyrrole compounds bearing beta-(5',6',7'-trimethoxy-2'-indolyl)acryloyl group

A series of A-ring pyrrole derivatives of duocarmycin bearing beta-(5',6',7'-trimethoxy-2'-indolyl)acryloyl group were synthesized, and evaluated for in vitro anticellular activity against HeLa S3 cells and in vivo antitumor activity against murine sarcoma 180 in mice. New Seg-B analogues bearing beta-(5',6',7'-trimethoxy-2'-indolyl)acryloyl group containing double bond as spacer had lower peripheral blood toxicity than the derivatives bearing 5',6',7'-trimethoxyindole-2'-carboxyl group in Seg-B of the natural type. Moreover, most of them exhibited potent antitumor activity against in vivo murine tumor models.     

Characterization of IL-2-dependent cytotoxic T-cell clones. III. Inhibition of killing activity by monosaccharides

The effects of monosaccharides on the cytotoxic activity of cytotoxic T lymphocytes (CTL) and three cloned long-term cytotoxic T-lymphocyte lines (CTLL) are compared. Uncultured CTL and clones CTLL-A2 and CTLL-A11 were derived from the peritoneal cavity of C57BL/6 mice immunized against the H-2Dd determinants on the BALB/c sarcoma Meth A. Clone CTLL-R5 was derived from spleen of (BALB/c X C57BL)F1 mice immunized against a unique determinant on the BALB/c radiation-induced leukemia RL male 1. The cell-surface phenotype of the clones is Lyt-1+,2+,3+. Cytotoxic activity of CTLL-A2 and CTLL-R5 as determined by a 4-hr 51Cr-release assay was inhibited over 50% by 1 mM 2-deoxy-D-glucose. CTLL-A11 and the uncultured cytotoxic T cells were more resistant to inhibition by 2DG (40% at 20 mM). Surprisingly, it was found that the addition of D-mannose, D-galactose, D-glucose, L-fucose, alpha-methyl-D-mannose, and N-acetyl-D-glucosamine also inhibited, in a dose-related manner, the cytotoxicity of CTLL-A2 and CTLL-A11. CTLL-R5 showed a more restricted inhibition pattern: only D-mannose and D-galactose were inhibitory. The mechanism of inhibition remains to be clarified.     

A novel antibiotic produced by <Emphasis Type="Italic">streptomyces noursei</Emphasis> Da07210

A novel antibiotic 210-A, named as (6S,8aS,9S,11S,12aR)-6-hydroxy-9,10-dimethyldecahydrobenzo[d]azecine-2,4,12(3H)-trione, was isolated from the fermentation broth of Streptomyces noursei Da07210, its structure was unambiguously established by spectral analyses and chemical comparison with related cycloheximide. Experiments demonstrated that 210-A bore strong activity against Fusarium oxysporum f. sp. cubense race four (Foc race four), which also showed antitumor activity against SMMC-7721 human hepatocarcinoma cells and S180 murine sarcoma, and the IC₅₀ values were 0.77 and 0.74 μg/ml, respectively.     

Bioassay-guided isolation of an alkaloid with antiangiogenic and antitumor activities from the extract of Fissistigma cavaleriei root

Fissistigma cavaleriei (Levl) Rehd (Annonaceae) is used as a folklore medicine for treatment of inflammation, arthritis, and tuberculosis by Miao people in China. In the present study, the antiangiogenic activity of F. cavaleriei was investigated. The chorioallantoic membrane of the fertilized hen's egg (CAM assay) was used to determine antiangiogenic activity of the plant extract. Compound (1), a compound with antiangiogenic activity, was isolated by bioassay-guided fractionation from F. cavaleriei for the first time. The structure of compound (1) was elucidated on the basis of spectroscopic methods. Colorimetric COX (ovine) inhibitor screening assay was used to determine its inhibitory effect on COX-1 and COX-2. MTT and Sulforhodamine B assays were used to investigate its cytotoxic effects on tumor cell lines. As a result, compound (1) showed a selectively inhibiting effect on COX-2 and could inhibit the growth of tumor cells in vitro. The antitumor activity of compound (1) was further confirmed by the observation that compound (1) administration significantly inhibited the growth of S-180 cells in mice. Moreover, compound (1) was able to enhance the antitumor activity of doxorubicin in the mice bearing with S-180 cells while combined with doxorubicin. In conclusion, compound (1) is a multi-target molecule and further experimental investigations are needed to determine whether it can be used as a lead molecule for tumor treatment.     

Synthesis and properties of the retro-analogue of myelopeptide MP-2

The bone marrow myelopeptide MP-2 (Leu-Val-Val-Tyr-Pro-Trp), exhibiting antitumor activity, and its retro-analogue (Trp-Pro-Tyr-Val-Val-Leu) were synthesized, and their properties were studied. The in vitro and in vivo activities of retro-MP-2 were comparable with those of MP-2. Both peptides equally restored the functional activity of T-lymphocytes inhibited by toxins released by HL-60 cells and inhibited by 70-82% the growth of various types of transplantable solid tumors: Ca-755 adenocarcinoma of the mammary gland, Lewis adenocarcinoma of the lung, and S180 sarcoma. The positions and intensities of the Cotton effects in CD spectra of the MP-2 peptide and its retro-analogue in various solvents are almost indistinguishable. The positions of extrema and integral intensities of the amide I and amide A bands in IR spectra of both peptides were practically identical. The English version of the paper: Russian Journal of Bioorganic Chemistry, 2005, vol. 31, no. 3; see also http://www.maik.ru.     

Synthesis and antitumor activity of duocarmycin derivatives: modification at C-8 position of A-ring pyrrole compounds bearing the simplified DNA-binding groups

A series of the 8-O-substituted A-ring pyrrole derivatives of duocarmycin bearing the simplified DNA-binding moieties such as cinnamoyl or heteroarylacryloyl groups were synthesized, and evaluated for in vitro anticellular activity against HeLa S3 cells and in vivo antitumor activity against murine sarcoma 180 in mice. In addition, the stability of the 8-O-substituted analogues in aqueous solution and the conversion to their active form (cyclopropane compound) from the 8-O-substituted analogues in mice or human serum were examined. The 8-O-substituted A-ring pyrrole derivatives bearing the simplified DNA-binding moieties showed remarkably potent in vivo antitumor activity and low peripheral blood toxicity compared with the 8-O-substituted A-ring pyrrole derivatives having the trimethoxyindole skeleton in segment-B (Seg-B), which were equal to 8-O-[(N-methylpiperazinyl)carbonyl] derivatives of 4'-methoxycinnamates and 4'-methoxy-beta-heteroarylacrylates. Moreover, among 8-O-substituted analogues, several compounds can be chemically or enzymatically converted to their active form in human serum. This result indicated that new 8-O-substituted derivatives were different prodrugs from KW-2189 and 8-O-substituted analogues being the same type of prodrug as KW-2189.     

Antitumor, genotoxicity and anticlastogenic activities of polysaccharide from Curcuma zedoaria

The antitumor effect of the partially purified polysaccharide from Curcuma zedoaria was studied in mice transplanted with sarcoma 180 cells. The polysaccharide fraction, CZ-1-III, at dose of 6.25 mg/kg/d showed 50% inhibition in solid tumor growth. When mice were injected with fractions, CZ-1 and CZ-1-III, at the dose of 100.0 mg/kg, 91.6% and 97.1% of tumor growth were inhibited, respectively, indicating that the cytotoxic effect of polysaccharide on sarcoma 180 cells increases upon increasing the amount of polysaccharide administered. To assess the genotoxicity of CZ-1-III fraction, several classical toxicological tests were performed. In Ames test, CZ-1-III did not show any transformation of revertant with or without S-9 metabolic activating system, indicating the lack of mutagenic effect of the compound. To assess clastogenic effect, micronucleus and chromosomal aberration assays were performed using Chinese hamster lung (CHL) fibroblast cells. However, up to 259.0 microg/ml concentration of CZ-1-III, neither micronucleus formation nor chromosomal aberration was induced regardless of the presence of S-9 metabolic activating system. Inhibition of CZ-1-III on micronucleus formation induced by mitomycin C was exhibited in a dose-dependent manner, maximally up to 52.0%. These results strongly suggest that CZ-1-III, the polysaccharide fraction from C. Zedoaria, decreases tumor size of mouse and prevents chromosomal mutation.     

Mechanism of target cell lysis by cytotoxic T lymphocytes (CTL) employing RSV-induced H-2 congenic and recombinant mouse tumor cells: demonstration of soluble mediators released from H-2-restricted CTL clones

The secretion and the specificity of cytotoxic mediators from H-2-restricted cytotoxic T lymphocytes (CTL) were examined using non-virus-producing target tumor cells induced by the Schmidt-Ruppin strain of Rous sarcoma virus (SR-RSV) in B10 congenic and recombinant mice. By using rat concanavalin A supernatant, two H-2-restricted CTL clones were established from cytotoxic effector cells of B10.A(5R) mice primed with SR-RSV-induced syngeneic tumor Cell-free supernatants from the H-2-restricted CTL clones cocultured with syngeneic tumor cells had selectively high cytotoxic activity for syngeneic and H-2-compatible tumor cells, but not for H-2-incompatible tumor cells. YAC-1 cells, and B10.A(5R) blasts as defined in the 5-hr 51Cr-release assay. The cytotoxic activity was detected in the cell-free supernatants from the CTL clones cocultured with the CTL-sensitive syngeneic and H-2-compatible tumor cells, but not with the CTL-insensitive tumor cells and YAC-1 cells. The cytotoxic activity of the cell-free supernatant could be adsorbed by the syngeneic tumor cells, but not by YAC-1 and L(s) cells. Thus, the H-2-restricted CTL clones against SR-RSV-induced tumor cells were capable of releasing cytotoxic mediators by coculturing with syngeneic or H-2-compatible tumor cells, and the cytotoxic mediators showed a certain H-2-restricted manner in killing the target cells. These results suggest that the lysis of RSV-induced tumor cells by H-2-restricted CTL can at least in part be mediated by cytotoxic factors.     

Purification and characterization of a novel cytotoxic substance from cell-free extract of Streptococcus pyogenes

A cytotoxic substance designated as streptococcal cytotoxic protein (SCP) was isolated from a cell-free extract of the Su strain of Streptococcus pyogenes possessing cytotoxic and antitumor activity. SCP was purified with a series of column chromatography and preparative PAGE to give a homogeneous single band as revealed by PAGE analysis. The purified SCP has a molecular mass of 165 kDa, composed of four 43 kDa subunits, and its pI is 4.3. SCP was sensitive to proteinases and was labile to heat and at acidic or alkaline pH. SCP showed inhibitory effects on the [3H]thymidine, [3H]uridine and [3H]leucine uptakes and on the growth of cells, and released 51Cr from cells when the protein was added to the cultures of Ehrlich ascites carcinoma (EAC), mouse mammary tumor (MM-2), leukemia (L-1210) and NIH-3T3 mammalian cells in vitro. SCP also showed an antitumor effect on EAC or MM-2 tumor-bearing mice but not on L-1210 tumor-bearing mice in vivo.