Structure toxicity relationships of synthetic azaspiracid-1 and analogs in mice

Synthetic azaspiracid-1 (AZA-1, 1), 6-, 10-, 13-, 14-, 16-, 17-, 19-, 20-epi-azaspiracid-1 (C₁–C₂₀-epi-AZA-1, 2), and twelve truncated azaspiracid-1 analogs (3–14) were synthesized and tested for their toxicity effects in mice. Of these compounds only AZA-1 (1) and its diastereomer C₁–C₂₀-epi-AZA-1 (2) exhibited significant toxicity in mice with the latter compound (2) being one-fourth as toxic as the former (1). The lack of toxicity exhibited by the severely truncated analogs (3–14) implies that the entire or at least a major part of the structure of AZA-1 (1) is required for biological activity.     

Comparison of biologic activities of synthetic lipopentapeptide analogs of bacterial lipoprotein in mice

Mitogenicity, lethal toxicity, induction of tumor necrotizing factor (TNF), and antitumor activity against Meth A fibrosarcoma of four chemically synthesized lipopentapeptide analogs, S-[2,3-bis(palmitoyloxy)-2R (designated as KAB-1), -2S(KAB-3)-propyl]-N-palmitoyl-(R)-cysteinyl-(S)-seryl- (S)-seryl-(S)-asparaginyl-(S)-alanine, S-[2,3-bis(palmitoyloxy)-2R(KAB-2), and -2S(KAB-4)-propyl]-N-[(2,2,2)-trichloroethoxycarbonyl]-(R)- cysteinyl-(S)-seryl-(S)-seryl-(S)-asparaginyl-(S)-alanine, of bacterial lipoprotein were investigated. These four analogs, as well as bacterial lipopolysaccharide (LPS) or synthetic Escherichia coli-type lipid A (506), were capable of increasing of [3H]thymidine into splenocytes of C3H/He mice. Although LPS and 506 did not exhibit the mitogenic activity in C3H/HeJ mice, KAB compounds showed remarkable mitogenicity. These analogs did not show the lethal toxicity at a high dose of 50 micrograms/mouse in galactosamine-loaded C57BL/6 mice. Peritoneal macrophages, stimulated with four analogs, caused the production of TNF which induces the L929 cell lysis in vitro. Twice, intravenous injections of 50 micrograms/mouse of these analogs showed weak growth inhibition of Meth A fibrosarcoma in BALB/c mice. The inhibitory effect of KAB-2 compound, which caused the strong TNF-induction among the four analogs, was the most potent. These results indicate that the biological activity of KAB-2 (R-configuration of the C-2 position in glycerol moiety with dipalmitoyl) is stronger than that of the other three analogs.     

Soybean pod set enhancement with synthetic cytokinin analogs

The previously reported activity of benzyladenine and selected other cytokinin analogs to increase pod set in soybean (Glycine max [L.] Merr.) was further investigated to define the structure-activity relationship and evaluate the effects of the cytokinins on yield parameters. Enhancement of pod set was found to be greatest with N-6 saturated alkyl substituted analogs, and was only weakly associated with activity in a callus growth bioassay. The response of yield parameters to increasing pod load was evaluated by applying various cytokinin analogs having a range of pod set enhancement activity. The increased pod load at the treated nodes was not compensated by a reduction in pod number on the remainder of the plant. However, there was a compensatory decrease in seed size. Overall, a significant trend to greater total seed weight per plant was associated with the increased pod number. Initial evaluations indicated that foliar applications of select cytokinins could temporarily increase pod number. However, the increases in pod number obtained with foliar treatments were too small to be of practical utility and were not maintained to maturity.     

Metabolism of synthetic inositol trisphosphate analogs

A series of synthetic analogs was employed to explore structure-activity relationships in the metabolism of the second messenger inositol trisphosphate (IP3) in vascular tissue. Cytosolic IP3-5-phosphatase activity was purified approximately 240-fold from bovine aorta. All synthetic analogs tested were apparent competitive inhibitors of the 5-phosphatase activity. The order of potency was DL-1,3,4,5-IP3 greater than D-1,4,5-IP3 greater than DL-1,3,4-IP3 greater than L-1,4,5-IP3 greater than 1,3,5-IP3 greater than DL-6-methoxy-1,4,5-IP3 greater than DL-2,4,5-IP3 greater than DL-1,2,4-cyclohexane-P3. The least potent analogs had Ki values only 11 times higher than the apparent Km of the substrate D-1,4,5-[3H]IP3. However, only three synthetic compounds, DL-1,3,4,5-IP4, D-1,4,5-IP3, and DL-2,4,5-IP3, could serve as substrates for the 5-phosphatase. IP3 kinase activity in the same tissue exhibited considerably more selectivity with respect to inhibition by IP3 analogs. D-1,4,5-IP3 was about 30 times more potent than DL-1,3,4,5-IP4 and 100-1000 times more potent than the other compounds tested. The function of the IP3 receptor was evaluated by measuring labeled calcium mobilization in permeabilized bovine aortic smooth muscle cells in culture. While all analogs tested were full agonists, vast differences in potency were observed. D-1,4,5-IP3 was about 30 times more potent than DL-2,4,5-IP3 and 100-2000 times more potent than the other analogs tested. The results suggest that IP3-5-phosphatase activity is relatively nonselective in the binding of inositol polyphosphates, while IP3 kinase activity and the IP3 receptor exhibit great selectivity in the recognition of these compounds.     

Protective effects of 5,6,7,8-tetrahydroneopterin against X-ray radiation injury in mice

The protective effects of 5,6,7,8-tetrahydroneopterin (NH4) against radiation injury in mice were studied. (C57BL/6xA/J)F1 (B6A) mice received a single whole-body irradiation dose of 200, 400, 700 or 800 cGy of X-rays. NH4 (30 mg/kg body weight) or phosphate-buffered saline (PBS) was injected intraperitoneally into irradiated mice 10 min before and after the irradiation and again after 6 h. All mice which received the 800 cGy radiation+PBS died between 8 and 11 days after the treatment. In contrast, those which also received NH4 demonstrated a significantly prolonged survival time and 40% lived more than 5 months. Total numbers of thymocytes and spleen cells on day 5 post-irradiation were dramatically reduced in line with the radiation dose. The survival was significantly enhanced by NH4 in treated mice. The proliferation of spleen cells in mice stimulated by concanavalin A (Con A) or lipopolysaccharide (LPS) was also greater in NH4 treated mice. The immune response of survivors 5 months after 800 cGy+NH4 treatments, against Con A, LPS, allogenic mouse, and sheep red blood cells had essentially recovered to the levels of normal mice. These results indicate that NH4 had an important role in modifying radiation injury.     

Comparative analysis of proinflammatory cytokines in plasma of mice exposed to radiation or in combined radiation injury

Male mice (CBA x C57BL/6)F1 were used for the experiments throughout this study. Blood serum levels of IL-1 beta, IL-6, TNF-alpha, IL-3, and GM-CSF were evaluated by means of enzyme-linked immunosorbent assay at 3, 6, 24, 48 and 72 hours after 7 Gy gamma-irradiation alone or combined injury (irradiation + thermal burn). Radiation as well as combined injury did not cause any important alterations of IL-1 beta, TNF-alpha, IL-3, and GM-CSF concentrations in the system circulation. Combined injury revealed more enhanced serum levels of IL-6 versus only irradiated mice. A possible significance of this phenomenon at the combined radiation and thermal burns' pathogenesis is discussed.     

Experimental basis of cancer combination chemotherapy with retinoids,cytokines, 1,25-dihydroxyvitamin D3, and analogs

Retinoids, cytokines as well as 1,25-dihydroxyvitamin D₃ [1,25(OH)₂D₃] and analogs possess properties known to contribute potentially to cancer chemopreventive and chemotherapeutic effects. They induce cell differentiation, inhibit cell prolifeation, suppress expression of viral oncogenes, and inhibit angiogenesis nexessary for tumor growth. Since clinical combination chemotherapy of cytotoxic agents has proven superior to monotherapy, This moddality might also be useful for other classes of antitumor drugs. A series of retinoids, such as all-trans-, 13-cis-, 9-cis retinoic acid, and acitreti, cytokines, 1,25(OH)₂D₃, and analogs have been investigated in mocdel systems of defferentiation, proliferation, viral oncogenes, and angiogenesis. The three classes of compounds have common effects but nevertheless show a variance depending on the particular representative of each class. Combination of compounds of the different classes led in the various models to a hegher efficacy compared with the compounds given alone. Cytokines such as IFNα, IFNγ, G-CSF, TNFα, IL-1, and IL-4 markedly potentiate the differentiation-inducing effect of retinoids. Cytokines as well as retinoids combined with 1,25(OH)₂D₃ and analogs synergistically enhanced differentiation induction in human transformed hemopoietic cell lines. On a series of human transformed epithelial cell lines a panel of cytokies, such as IFNα, IFNβ, TNFα, TGFβ, and EGF acted synergistically with retinoies on inhibition of proliferation. This was also observed by combining retinoids with 1,25(OH)₂D₃ and analogs. Retinoids as well as interferons α and γ have the capacity to suppress the capacity to supperess the oncogene expression of human papilloma viruses which are involved in induction and growth of certain malignancies such as cervical cancer. All-trans-, 13-cis-, 9-cis retinoic acid, and acitretin as well as IFNα inhinited the formation of newly formed blood vessels induced by injection of HPV harboring, tumorigenic cell lines into Balb/C mice. The combination of retinoids with IFNα was more efficacious in inhibition angiogenesis than a retinoid or IFNα given alone. Since there is evidence that cell differentiationl, cell proliferation, viral oncogene expression as well as angiogenesis play a role in tumor induction and/or progression of tumor growth, retinoids, cytokines, as well as 1,25(OH)₂D₃ and analogs may be useful in chemoprevention and chemotherapy of neoplasms. based on the superior effect of combinations compared with administration of single compounds of these three classes under experimental conditions there is hope that also clinical application of combination treatment might bring as a step forward in chemopreventio and chemotherapy of cancer. This has already been proven in a very limited number of very limited numberr of clinical trials.     

In vivo roles of lysophospholipid receptors revealed by gene targeting studies in mice

Lysophosphatidic acid (LPA) and sphingosine 1-phosphate (S1P) are extracellular ligands for a family of G protein-coupled receptors (GPCRs), LPA1/2/3 and S1P1/2/3/4/5. Through coupling to multiple classes of G proteins and activating multiple signaling pathways, LPA/S1P receptors have been shown to be integral players for many essential cellular and physiological processes. Generation and analysis of mice deficient in each of LPA1, LPA2, S1P1, S1P2, and S1P3 have provided valuable information on the in vivo roles of these receptors. This review is focussed on expression patterns of each receptor gene in wild-type mice, targeted deletion approaches for generating mutant animals, main phenotypes of receptor-null mice, and alterations in signaling characteristics in receptor-deficient primary cells. Altogether, these data give insights to the importance of LPA/S1P receptors at the cellular and organismal level.     

Effect of the radioprotector adeturon on radiation injury in mice with Lewis carcinoma of the lung

A study was made of the radioprotective effect of Adeturon, a protective agent obtained in Bulgaria, on mice with Luis lung tumors exposed to fractionated radiation. The effect of the radioprotector on a radiation-induced injury to normal tissue was estimated by the number of leucocytes in the peripheral blood and its count, cellularity of bone marrow and spleen and the mass of the latter, and by the number of exogenous and endogenous CFUs. A pronounced radioprotective effect of Adeturon was implemented through maintaining or normalizing the indices under study impaired by tumor inoculation or irradiation.     

Experimental study of combined effect of actoprotectors and radioprotectors in radiation injury

Combined treatment of mice and dogs with actoprotector bimethyl and radioprotector indralin suppressed radioprotective action of the latter, the surveillance being decreased from 83.3 to 14.3% in dogs and from 70.3 to 5% in mice. At the same time vasodynamic action of indralin (increase of artherial pressure up to 30% from a reference level) did not change.     

Antitumor activity against Meth A fibrosarcoma and biologic activities of synthetic monosaccharide analogs of lipid A in mice

Antitumor activity, mitogenicity, and lethal toxicity of chemically synthesized lipid A analogs, acylglucosamine-4- or -6-phosphate with the alpha, beta-hydroxyacyl, acyloxyacyl, or hydroxyacyloxacyl groups at the C-2 and C-3 positions, were examined. Meth A fibrosarcoma cells (5 X 10(5)) were inoculated subcutaneously into BALB/c mice on day 0, and six compounds (50 micrograms/mouse) were administered intravenously on days 7 and 9. Although the antitumor activity of these compounds was weaker than that of natural lipopolysaccharide (LPS) or the synthetic lipid A analog (506) of Escherichia sp type, all groups exhibited tumor inhibition rates of 40% to 50% and delayed tumor growth. Six compounds, with the exception of compound A-173 (with the hydroxytetranoyl group at the C-2 and C-3 positions), were capable of increasing the incorporation of [3H]thymidine into cultured splenocytes of C57BL/6 mice, and caused lethal toxicity in C57BL/6 mice sensitized with galactosamine. However, these compounds had lower toxicity than bacterial LPS (about 500- to 1,000-fold). Compounds A-172 and A-174, which have the same structure except for the C-4 or C-6 position of the phosphate group, exerted similar antitumor activity, mitogenicity, and lethality. The results discussed above indicate that the biologic activity of these compounds correlates with the carbon number of fatty acid but is not affected by the different location of the phosphate group. Furthermore, it seems that the difference between the alpha, beta-hydroxy position of fatty acid and the R or S configuration does not alter the biologic effects.     

Radioprotective activity of Polyalthia longifolia standardized extract against X-ray radiation injury in mice

The radioprotective effect of Polyalthia longifolia was studied in mice. P. longifolia treatment showed improvement in mice survival compared to 100% mortality in the irradiated mice. Significant increases in hemoglobin concentration, and red blood cell, white blood cell and platelet counts were observed in the animals pretreated with leaf extract. Pre-irradiation administration of P. longifolia leaf extract also increased the CFU counts of the spleen colony and increased the relative spleen size. A dose-dependent decrease in lipid peroxidation levels was observed in the animals pretreated with P. longifolia. However, although the animals pretreated with P. longifolia exhibited a significant increase in superoxide dismutase and catalase activity, the values remained below normal in both liver and the intestine. Pre-irradiation administration of P. longifolia also resulted in the regeneration of the mucosal crypts and villi of the intestine. Moreover, pretreatment with P. longifolia leaf extract also showed restoration of the normal liver cell structure and a significant reduction in the elevated levels of ALT, AST and bilirubin. These results suggested the radioprotective ability of P. longifolia leaf extract, which is significant for future investigation for human applications in developing efficient, economically viable, non-toxic natural and clinically acceptable novel radioprotectors.     

Conformation states of thymopentin and its synthetic analogs

Spatial structures of the biologically active fragment Arg-Lys-Asp-Val-Tyr of tymopoetin (tymopentin) and its synthetic analogs: [Lys1-Arg2]-, [Glu3]- and [Gly3] have been investigated by theoretical conformational analysis. These results indicate that the conformational properties of fragments are represented in aqueous solution by five different backbone forms.