Synthesis and anti-tumor-promoting activity of glycoglycerolipid analogues lacking the glycerol backbone

Glycoglycerolipid analogues lacking the glycerol backbone were prepared through a lipase catalyzed transesterification of beta-D-galactosylethylene glycol. The inhibitory effect of the resultant isomeric hexanoates at the primary alcoholic positions, beta-D-galactosylethylene glycol itself and nonyl beta-D-galactopyranoside, was tested on Epstein-Barr virus early antigen (EBV-EA) activation in Raji cells promoted by the tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA), as a primary screening test for inhibitors of tumor promotion. All the compounds assayed were found to be less active than the reference 2-O-beta-D-galactopyranosylglycerol derivatives, of which they are simplified models, indicating that the anti-tumor-promoting activity is very closely related to the presence of a free hydroxymethylene group on the glycerol-like aglycone moiety.     

Effect of some benzyl alcohols on rooting of bean cuttings

The root differentiating properties of various benzyl alcohols were studied using a bean rooting test. The results showed that the methoxy derivatives enhanced rooting more efficiently than did the hydroxy derivatives. Only 4-hydroxybenzyl alcohol inhibited rooting, but the addition of IAA at 10^?5M nullified the inhibition. Root promotion by the alcohols with a hydroxy group in the o-or p-position was increased by the addition of IAA. Auxin did not modify the rooting stimulation caused by the methoxy derivatives. The position of the methoxy group did not influence the rooting activity.     

Chronic exposure to ionizing radiation as a tumor promoter in mouse skin.

We have tested chronic exposure to 90Y beta radiation for its action as a complete tumor promoter, a stage I tumor promoter, or a stage II tumor promoter in SENCAR mouse skin. In skin initiated with a single application of 7,12,dimethylbenz[a]anthracene (DMBA, 10 nmol), chronic exposure to beta radiation as a complete promoter (0.5 Gy, twice/week, 13 weeks) produced no tumors and, when added to a complete chemical promoter (TPA), reduced tumor frequency about 30%. A similar result was observed when beta radiation was tested as a stage II promoter. DMBA-initiated mice that received chemical (12-O-tetradecanoylphorbol-13-acetate, TPA) stage I promotion followed by 13 weeks of beta-radiation exposure (0.5 Gy, twice/week) as stage II promotion produced essentially no tumors, and combining the same chronic beta-radiation exposure with chemical (mezerein) stage II promotion reduced tumor frequency about 20% when compared to a similar group that was not irradiated. Chronic beta-radiation exposure was tested two ways as a stage I tumor promoter in initiated skin that was subsequently treated with mezerein as a stage II promoter. Stage I promotion was shown to proceed with the passage of time, indicating this process occurs naturally in the absence of chemical or physical stimulation. Hyperthermia, previously shown to be a potent inhibitor of chemically stimulated stage I promotion, had no effect on the natural process, indicating at least some differences in mechanism between the two processes. The natural process was, in fact, inhibited by chemical tumor promoters, but not by radiation. In addition to the increase resulting from this natural process, tumor frequency was further increased slightly but significantly (12-15%, P less than or equal to 0.05) when chronic radiation exposure was given as a stage I promoter (0.5 Gy, twice/week, 13 weeks) subsequent to initiation, in spite of the expected 20% reduction resulting from this dose. Exposure of initiated animals to radiation (0.5 or 1.0 Gy, twice/week, 2 weeks) in addition to TPA as stage I promotion produced a similar increase in tumor frequency (P less than 0.02). At higher radiation doses, however, tumor frequency was reduced compared to unirradiated controls. In a third test as a stage I promoter, beta radiation (0.5 Gy twice/week, 4 weeks) was given prior to initiation with N-methyl-N'-nitro-N-nitrosoguanidine in animals subsequently promoted by TPA (twice/week, 13 weeks), and again the radiation slightly but significantly (P less than 0.03) increased tumor frequency compared to the unirradiated control group.(ABSTRACT TRUNCATED AT 400 WORDS)     

Protein kinase C as a possible receptor protein of tumor-promoting phorbol esters

A tumor-promoting phorbol ester, [3H]phorbol-12,13-dibutyrate, may bind to a homogeneous preparation of Ca2+-activated, phospholipid-dependent protein kinase (protein kinase C) in the simultaneous presence of Ca2+ and phospholipid. This tumor promoter does not bind simply to phospholipid nor to the enzyme per se irrespective of the presence and absence of Ca2+. All four components mentioned above appear to be bound together, and the quaternary complex thus produced is enzymatically fully active for protein phosphorylation. Phosphatidylserine is most effective. Various other phorbol derivatives which are active in tumor promotion compete with [3H]phorbol-12,13-dibutyrate for the binding, and an apparent dissociation binding constant of the tumor promoter is 8 nM. This value is identical with the activation constant for protein kinase C and remarkably similar to the dissociation binding constant that is described for intact cell surface receptors. The binding of the phorbol ester is prevented specifically by the addition of diacylglycerol, which serves as activator of protein kinase C under physiological conditions. Scatchard analysis suggests that one molecule of the tumor promoter may bind to every molecule of protein kinase C in the presence of Ca2+ and excess phospholipid. It is suggestive that protein kinase C is a phorbol ester-receptive protein, and the results presented seem to provide clues for clarifying the mechanism of tumor promotion.     

青蒿素及其衍生物的抗肿瘤机制

恶性肿瘤是严重威胁人类健康的重大疾病。尽管治疗手段不断发展,但推广度及疗效仍极为有限。新近研究发现,经典抗疟一线药青蒿素及其衍生物具有广泛抗肿瘤活性。大量研究提示,青蒿素及其衍生物通过细胞毒性效应直接杀死肿瘤细胞,也可诱导细胞周期阻滞从而抑制细胞增殖。另一方面,可通过凋亡、自噬、铁死亡途径导致细胞死亡。还可调控肿瘤微环境,从而抑制肿瘤细胞侵袭与转移。然而,尽管青蒿素及其衍生物展现出强大的抗肿瘤潜能,但其作用机制仍十分复杂。本文就青蒿素及其衍生物的抗肿瘤机制及其研究进展作一综述。     

恶性肿瘤胸腹水细胞学检验的探讨

目的探讨胸腹水细胞学检验在恶性肿瘤细胞诊断中的应用价值。方法回顾我院2014年1月~2016年6月376例胸腹水脱落细胞学的检验结果,分析胸腹水细胞学检验在恶性肿瘤诊断中的应用价值。结果 376例胸腹水脱落细胞学检验患者中,发现疑似肿瘤细胞3例,恶性肿瘤细胞74例,病理组织学证实恶性肿瘤患者79例,胸腹水细胞学检验的正确率高达97.47%,假阴性率为2.53%,假阳性率为0%。结论胸腹水脱落细胞学检验在恶性肿瘤诊断中具有重要的临床价值,值得临床推广。     

Design,Synthesis and Antitumor Activity of FAK/PLK1 Dual Inhibitors with Quinazolinone as the Skeleton

Febrifugine is a kind of quinazolinone compound with high biological activity from a Chinese herb called Chang Shan (Dichroa febrifuga). Febrifugine and its derivatives possess extensive biological activities, some of which exhibited anti-tumor activities as FAK inhibitors. However, they are not very effective at inhibiting tumor metastasis, perhaps because tumors gain energy through compensatory activation of other signaling pathways that promote cell migration and invasion. Therefore, seventeen novel febrifugine derivatives with quinazolinone skeleton were designed, synthesized and acted as potential FAK/PLK1 dual inhibitors. These compounds were determined by ¹H-NMR, ¹³C-NMR and MS. Most of the compounds exhibited good inhibitory activity against cancer cell lines by computer-assisted screening, antitumor activity test and FAK/PLK1 inhibitory activity test, wherein compound 3b was screened as a high-efficiency lead compound.     

Detection of initiating as well as promoting activity of chemicals by a novel cell transformation assay using v-Ha-ras-transfected BALB/c 3T3 cells (Bhas 42 cells)

Cell transformation assay using BALB/c 3T3 cells, C3H10T1/2 cells and others, can simulate the two-stage carcinogenesis utilized for formation of transformed foci. A sensitive cell transformation assay for tumor initiators as well as promoters has been developed using a v-Ha-ras-transfected BALB/c 3T3 cell line, Bhas 42; these cells are regarded as initiated in the two-stage paradigm of carcinogenesis. To distinguish between initiation and promotion, the initiation assay involves a 2-day treatment of low-density cells, obtained one day after plating, with a test chemical, and the promotion assay involves treatment of near-confluent cells with a test chemical for a period of 12 days (Day 3-14). When Bhas 42 cells were treated with tumor initiators, N-methyl-N'-nitro-N-nitrosoguanidine and 3-methylcholanthrene, transformed foci were induced in the initiation assay but not in the promotion assay. In contrast, tumor promoters, 12-O-tetradecanoylphorbol-13-acetate, lithocholic acid and okadaic acid, gave negative responses in the initiation assay but positive responses in the promotion assay. The results were reproducible with various treatment protocols. Sixteen polycyclic aromatic hydrocarbons were examined using both assays. Benzo[a]pyrene and 7,12-dimethylbenz[a]anthracene induced focus formation only in the initiation assay. Increase of focus formation was observed in the promotion assay with benzo[e]pyrene, benzo[ghi]perylene, 1-nitropyrene and pyrene. Benz[a]anthracene, benz[b]anthracene, chrysene and perylene showed positive responses in both initiation and promotion assays. Results of initiation and promotion assays of acenaphthylene, anthracene, coronene, 9,10-diphenylanthracene, naphthalene and phenanthrene were negative or equivocal. The present Bhas assays for the detection of either/both initiating and promoting activities of chemicals are sensitive and of high performance compared with other cell transformation assays.     

Topically applied vitamin E prevents massive cutaneous inflammatory and oxidative stress responses induced by double application of 12-O-tetradecanoylphorbol-13-acetate (TPA) in mice

Vitamin E (alpha-tocopherol) is a promising chemopreventive and pharmacologically safe agent, which can be exploited or tested against skin cancer. It is an established antioxidant with an ability to ameliorate the UV-induced skin damage and chemically induced inflammation in lungs. However, there are some conflicting reports about its role as a modulator of chemically induced promotion. We evaluated its efficacy in preventing the inflammatory and oxidative stress responses in a double 12-O-tetradecanoylphorbol-13-acetate (TPA) application tumor skin promotion protocol. Double application of TPA was undertaken to produce massive inflammatory and oxidative stress responses. Topical TPA treatment adversely altered many of the marker responses of stage I skin tumor promotion. Vitamin E application 30 min prior to TPA treatment (10 nmol) inhibited induction of hydrogen peroxide, myeloperoxidase (MPO) activity, xanthine oxidase (XO) activity and lipid peroxidation (LPO). Vitamin E also positively modulated altered antioxidants of mouse skin. Histological examination also revealed marked improvement. These results confirm the efficacy of vitamin E against early inflammatory and oxidative stress responses, which are hallmark of tumor promotion and provide rational basis for chemopreventive action of vitamin E in skin cancer.     

Proliferation and motility of HaCaT keratinocyte derivatives is enhanced by fibroblast nemosis

The role of paracrine tumor-stroma regulation in the progression of cancer is under intense investigation. Activated fibroblasts are key components of the tumor microenvironment providing the soluble factors mediating the regulation. Nemosis is an experimental model to study these parameters: formation of a multicellular spheroid activates fibroblasts and leads to increased production of soluble factors involved in the promotion of growth and motility. Role of nemosis was investigated in the tumorigenesis of HaCaT derivatives representing skin carcinoma progression. Conditioned medium from fibroblast spheroids increased proliferation rate of HaCaT derivatives. Expression of proliferation marker Ki-67 increased significantly in benign A5 and low-grade malignant II-4 cells, but did not further increase in the metastatic RT3 cells. Expression of p63, keratinocyte stem cell marker linked to cancer progression, was augmented by medium from nemotic fibroblasts; this increase was also seen in RT3 cells. Scratch-wound healing of the keratinocytes was enhanced in response to fibroblast nemosis. Neutralizing antibodies against growth factors inhibited wound healing to some extent; the response varied between benign and malignant keratinocytes. Migration and invasion were enhanced by conditioned medium from nemotic fibroblasts in benign and low-grade malignant cells. RT3 keratinocyte migration was further augmented, but invasion was not, indicating their intrinsic capacity to invade. Our data demonstrate that fibroblast nemosis increases proliferation and motility of HaCaT keratinocyte derivatives, and thus nemosis can be used as a model to study the role of soluble factors secreted by fibroblasts in tumor progression.     

Only a subset of 12-O-tetradecanoylphorbol-13-acetate-promoted mouse skin papillomas are promotable by benzoyl peroxide

The two-stage skin carcinogenesis model of initiation and promotion in Carcinogenesis-susceptible (Car-S) mice has been used to investigate the pathways of promotional activity of 12-O-tetradecanoylphorbol-13-acetate (TPA), a phorbol ester tumor promoter, and benzoyl peroxide (BzPo), a free radical-generating compound. To test whether distinct populations of 9,10-dimethyl-1,2-benzanthracene (DMBA)-initiated epidermal keratinocytes are responsive to the two promoters, tandem experiments were performed. DMBA-initiated Car-S mice were promoted twice weekly with maximal promoting doses of TPA or BzPo. When the number of papillomas/mouse reached a plateau, promotion in the TPA and BzPo groups was switched to BzPo or TPA, respectively, until achievement of a new plateau. Mice promoted with BzPo developed 11.0 +/- 1.3 papillomas/mouse and subsequent TPA promotion induced 13.8 additional papillomas, for a total of 24.8 +/- 2.1 papillomas/mouse. TPA-promoted mice developed 23.3 +/- 1.1 papillomas/mouse, and subsequent BzPo promotion for 91 days did not promote additional papillomas. Our results show a less than additive tumor response after sequential promotion with BzPo and TPA, or vice versa, indicating that the pathways of promotional activity of TPA and BzPo are interacting. While the final papilloma yield was similar at the end of the two tandem promotion experiments independently of promoter sequence, the percentage of mice developing carcinomas was significantly higher in mice that were promoted with BzPo in the first stage. No significant differences in the frequency and type of c-Ha-ras mutations were observed in TPA- and BzPo-promoted tumors, suggesting that promotion of DMBA-initiated cells by BzPo requires introduction of additional molecular alterations compared to TPA.     

Understanding the dual nature of CD44 in breast cancer progression

CD44 has been the subject of extensive research for more than 3 decades because of its role in breast cancer, in addition to many physiological processes, but interestingly, conflicting data implicate CD44 in both tumor suppression and tumor promotion. CD44 has been shown to promote protumorigenic signaling and advance the metastatic cascade. On the other hand, CD44 has been shown to suppress growth and metastasis. Histopathological studies of human breast cancer have correlated CD44 expression with both favorable and unfavorable clinical outcomes. In recent years, CD44 has garnered significant attention because of its utility as a stem cell marker and has surfaced as a potential therapeutic target, necessitating a greater understanding of CD44 in breast cancer. In this review, we attempt to unify the literature implicating CD44 in both tumor promotion and suppression, and explain its dualistic nature.     

Synthesis and biological activities of high affinity taxane-based fluorescent probes

Three fluorescent probes 3a, 3b, and 4 have been synthesized through conjugation of fluorescein and difluorescein groups to the 7-OH of C-2 modified paclitaxel and cephalomannine derivatives with very high affinity to microtubules. All these probes exhibited potent tubulin assembly promotion and tumor cell killing activities, thus may be useful as tools for the determination of thermodynamic parameters and exploration of ligand–microtubule interactions.     

Down-regulation of particulate protein kinase Cepsilon and up-regulation of nuclear activator protein-1 DNA binding in liver following in vivo exposure of B6C3F1 male mice to heptachlor epoxide

The effects of in vivo administration of the cyclodiene tumor promoter heptachlor epoxide on mouse liver protein kinase C were studied in male B6C3F1 mice by protein kinase C activity assays and Western blotting under conditions known to increase the incidence of hepatocellular carcinoma because protein kinase C is thought to be critical in phorbol ester-induced tumor promotion. Under these test conditions, 20 ppm dietary heptachlor epoxide for 1-20 days increased cytosolic and decreased particulate total protein kinase C activities, while 10 ppm had no effect. Further, total cytosolic and particulate protein kinase C activities were decreased within 1 hour by 10 mg/kg intraperitoneal (i.p.) heptachlor epoxide. Western blotting showed that conventional protein kinase Calpha and beta isoforms were unaffected by heptachlor epoxide. Particulate novel protein kinase Cepsilon, however, was selectively down-regulated by 1, 10, and 20 ppm dietary heptachlor epoxide, whereas the cytosolic isoform was decreased by 1 and 10 ppm heptachlor epoxide for 10 days. The high-dose treatment for 24 hours also decreased particulate novel protein kinase Cepsilon but increased the cytosolic titer. These results demonstrate that this isoform is unique in its sensitivity to heptachlor epoxide. Activator protein-1 DNA binding, a critical factor in tumor promotion, was substantially increased at 3 and 6 hours with 3.7 mg/kg (i.p.) heptachlor epoxide and at 3 and 10 days with 20 ppm dietary heptachlor epoxide. The effects of heptachlor epoxide on protein kinase C and activator protein-1 are similar to those caused by phorbol ester treatments and correlate well to heptachlor levels found to induce tumors in mice. However, heptachlor epoxide did not initially activate protein kinase C with in vivo treatments or with in vitro treatments of a plasma membrane fraction aimed at demonstrating direct activation, as has been shown for phorbol esters. The ability of heptachlor epoxide to down-regulate particulate novel protein kinase Cepsilon correlates to dosages used in in vivo tumor promotion studies. However, this may represent a negative feedback response rather than a causative effect.     

Synthesis of amphiphilic derivatives of rose bengal and their tumor accumulation

It is known that the combination of laser light and its sensitizer is effective for noninvasive tumor treatment, referred to as photodynamic therapy. Using the combination of ultrasound and its sensitizer has also been suggested for a similar kind of tumor treatment, referred to as sonodynamic therapy. The purpose of this paper is to obtain such sensitizers accumulating selectively in tumors. Amphiphilic derivatives of rose bengal (RB) were synthesized to add a tumor-accumulating property to RB. One type of the synthesized RB derivatives (RBD3), having an alkyl chain with a branching carboxyl group, was found to be superior in amphiphilicity to the other types. Tumor tissue distribution of the synthesized derivatives in mice bearing colon 26 carcinoma was evaluated. It was found that RBD3s with carbon chain lengths of 12, 14, and 16 had higher concentrations in the tumor tissue than RB by more than 1 order of magnitude, several hours after administration. The concentrations correlated well with their water/1-octanol partition coefficients. Since RB is known to induce in vitro cell damage in combination with either laser light or ultrasound, the newly synthesized amphiphilic RB derivatives may be potentially useful as a tumor-selective sensitizer for both light and ultrasound.     

Glycyrrhizin exhibits potential chemopreventive activity on 12-O-tetradecanoyl phorbol-13-acetate-induced cutaneous oxidative stress and tumor promotion in Swiss albino mice

Glycyrrhizin and its aglycone, glycyrrhetic acid has been found useful for various therapeutic purposes. Glycyrrhizin has been shown to possess many physiological functions like anti-inflammatory activity, detoxification and inhibition of carcinogenic promoters. 12-O-Tetradecanoyl phorbol-13-acetate (TPA), a well-known phorbal ester is known for its tumor promotion activity. The induction of inflammation in skin mediated by TPA is believed to be governed by cyclooxygenase (COX), lipoxygenase and ornithine decarboxylase (ODC). These markers of inflammatory responses are important for skin tumor promotion. In our present study, we studied the chemopreventive effect of glycyrrhizin on TPA (20 nmol/0.2 mL acetone/animal, topically)-induced oxidative stress and hyperproliferation markers in skin. TPA enhanced lipid peroxidation with reduction in the level of catalase, glutathione, glutathione peroxidase, glutathione reductase and glutathione-s-transferase. TPA treatment also enhanced ODC activity and [3H] thymidine incorporation into cutaneous DNA. Prophylactic treatment of mice with glycyrrhizin (2.0 & 4.0 mg/0.2 mL acetone/animal, topically) resulted in a significant decrease in cutaneous microsomal lipid peroxidation (P < 0.001) and recovery of cutaneous glutathione content (P < 0.001) and its dependent enzymes. A significant inhibition in ODC activity and DNA synthesis (P < 0.001) was also observed. Thus, the results demonstrate that pretreatment with glycyrrhizin is protective against TPA-induced oxidative stress and tumor promotion in Swiss albino mice.     

Tamarix gallica ameliorates thioacetamide-induced hepatic oxidative stress and hyperproliferative response in Wistar rats

Tamarix gallica, a hepatic stimulant and tonic, was examined for its ability to inhibit thioacetamide (TAA)-induced hepatic oxidative stress, toxicity and early tumor promotion response in male Wistar rats. TAA (6.6 mmol/kg body wt. i.p) enhanced lipid peroxidation, hydrogen peroxide content, glutathione S-transferase and xanthine oxidase with reduction in the activities of hepatic antioxidant enzymes viz., glutathione peroxidase, superoxide dismutase and caused depletion in the level of hepatic glutathione content. A marked increase in liver damage markers was also observed. TAA treatment also enhanced tumor promotion markers, ornithine decarboxylase (ODC) activity and [3H] thymidine incorporation into hepatic DNA. Pretreatment of rats orally with Tamarix gallica extract (25 and 50 mg/kg body weight) prevented TAA-promoted oxidative stress and toxicity. Prophylaxis with Tamarix gallica significantly reduced the susceptibility of the hepatic microsomal membrane for iron-ascorbate induced lipid peroxidation, H2O2 content, glutathione S-transferase and xanthine oxidase activities. There was also reversal of the elevated levels of liver marker parameters and tumor promotion markers. Our data suggests that Tamarix gallica is a potent chemopreventive agent and may suppress TAA-mediated hepatic oxidative stress, toxicity, and tumor promotion response in rats.     

Recent studies on the biofunctions and biotransformations of curcumin

Curcumin is a major component of Curcuma species, which is commonly used as a yellow coloring and flavoring agent in foods. Curcumin has shown anti-carcinogenic activity in animals as indicated by its ability to block colon tumor initiation by azoxymethane and skin tumor promotion induced by phorbol ester TPA. Curcumin possesses anti-inflammatory activity and is a potent inhibitor of reactive oxygen-generating enzymes such as lipoxygenase/cyclooxygenase, xanthine dehydrogenase/oxidase and inducible nitric oxide synthase. Curcumin is also a potent inhibitor of protein kinase C, EGF-receptor tyrosine kinase and IkappaB kinase. Subsequently, curcumin inhibits the activation of NFkappaB and the expressions of c-jun, c-fos, c-myc and iNOS. It is proposed that curcumin may suppress tumor promotion through blocking signal transduction pathways in the target cells. Curcumin was first biotransformed to dihydrocurcumin and tetrahydrocurcumin and that these compounds subsequently were converted to monoglucuronide conjugates. These results suggest that curcumin-glucuronide, dihydro-curcumin-glucuronide, tetrahydrocurcumin-glucuronide and tetrahydrocurcumin are major metabolites of curcumin in mice.     

Possible role of DT-diaphorase in the bioactivation of antitumor quinones

Menadione derivatives that are toxic to tumor cells are believed to be reduced intracellularly to species that react with DNA. In this communication, we report evidence that one of these derivatives, 3-bromomethylmenadione, is reduced by DT-diaphorases present in rat liver cytosol and in rat 9L brain tumor cells. Dicoumarol, an inhibitor of DT-diaphorases was found to inhibit both the reduction of 3-bromomethylmenadione and its mutagenicity to Salmonella typhimurium TA 97. Homogenates of rat 9L cells were found to contain relatively high levels of DT-diaphorase, suggesting that these tumor cells may be relatively sensitive to antitumor quinones that are activated by this enzyme.