Effect of prostaglandin analogs: Latanoprost, bimatoprost, and unoprostone on matrix metalloproteinases and their inhibitors in human trabecular meshwork endothelial cells.

Bimatoprost, latanoprost, and unoprostone are prostaglandin F2α analogs (PGAs) and are used to lower intraocular pressure. We investigated the free acid effects of these three prostaglandin analogs: bimatoprost, latanoprost, and unoprostone on human matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMP) in the trabecular meshwork (TM) cells. Immunoblot results show that all three PGAs generally increased MMPs-1,9 and TIMPs-4. Additionally, bimatoprost and latanoprost both increased MMP-3 and TIMP-2, while unoprostone had an indeterminate effect on both. Zymography results show that all three PGAs except unoprostone increased intermediate MMP-1 activity while bimatoprost and latanoprost increased MMP-9 activity. Together, these data suggest that the balance between MMPs and TIMPs correlate to the relative intraocular pressure lowering effectiveness observed in clinical studies of these PGAs.

Ocular Pharmacology.

Medications are a common treatment for glaucoma. Pharmacologic agents include sympathomimetics, beta-blockers, miotics (direct acting and cholinesterase inhibitors), carbonic anhydrase inhibitors, and prostaglandin agonists. Other agents include hyperosmotic agents and nonsteroidal anti-inflammatory drugs. Members of the ophthalmic health-care team, with heightened attention to patient safety, prescribe, administer, monitor side effects, and educate patients on glaucoma medications.

Synthesis and in vitro cytotoxicity of cross-conjugated prostaglandin A and J series and their hydroxy derivatives.

The synthesis of two cross-conjugated prostaglandin analogues of known neurotrophic activity and their new hydroxy derivatives was accomplished starting from the diastereoisomeric (+)-camphor protected 3-[(dimethoxyphosphoryl)methyl]-4,5-dihydroxycyclopent-2-enones. The cytotoxicity of these compounds was determined against HeLa, K562, HL-60 human cancer cell lines and normal human cells (HUVEC). We found that NEPP11 and its C7-hydroxy derivative demonstrated high anticancer activity against the HeLa and HL-60 human cancer cell lines at concentrations ranging from 1 to 2 µM. Moreover, the C7-hydroxy derivative of NEPP11 displayed high cytotoxic selectivity between cancer cell lines and normal human cells. On the other hand, the J-type prostaglandin analogue of NEPP11 and its C13-hydroxy derivatives were much less toxic or nontoxic against the cancer and normal cells at concentrations up to 1 mM.

Traceless stereoinduction for the enantiopure synthesis of substituted-2-cyclopentenones.

The pseudoenantiomeric 4-O-Boc- and 4-OPMP-cyclopent-2-enones, readily available from hydroxymethylenefurane on multigram scale, are demonstrated to be exceptional building blocks for the synthesis of enantiopure 4-alkyl-5-(1'-hydroxyalkyl) substituted 2-cyclopentenones and derivatives thereof. The 4-OR substituent acts as a traceless stereoinducing element, conferring not only 1,2- but also 1,4-stereocontrol with excellent selectivity. The methodology developed here was applied for the rapid synthesis of natural products and biologically active 2-cyclopentenones such as TEI-9826, guaianes, and pseudoguaianolides.

O-Monoacyltartaric acid catalyzed enantioselective conjugate addition of a boronic acid to dienones: application to the synthesis of optically active cyclopentenones.

Enantioselective conjugate addition of styrylboronic acid to dienones was effectively catalyzed by an O-monoacyltartaric acid to afford monostyrylated products with good enantioselectivity. The RCM of the monostyrylated products using the Hoveyda-Grubbs II catalyst afforded optically active cyclopentenones, including a synthetic intermediate of the antitumor agent TEI-9826. The study shows that a diene additive such as 1,6-heptadiene or diallyl ether was essential for the RCM.

Enantioselective synthesis of 12-amino alkylidenecyclopentenone prostaglandins.

An enantioselective synthesis of new 12-amino alkylidenecyclopentenone prostaglandins is reported. The key step of the synthesis involved a [3.3] sigmatropic rearrangement of an asymmetric allylic cyanate to elaborate an asymmetric 5-amino-1,6-diene which was further transformed into cyclopentenone by successive ring-closing metathesis reaction catalyzed by the Grubbs reagent and one-pot oxidation. A palladium-catalyzed cross-coupling reaction on a 5-iodo-1,5-diene allowed the synthesis of prostanoids with variable Rw side chains. These new compounds exhibit high cytotoxic activities.

Anti-cancer-prostaglandin-induced cell-cycle arrest and its modulation by an inhibitor of the ATP-dependent glutathione S-conjugate export pump (GS-X pump).

The A and J series of prostaglandins (PGs) accumulate in the nuclei to suppress the proliferation of cancer cells. Here we report that Delta7-PGA1 methyl ester, a synthetic anti-cancer PG, increased the level of mRNA for the cyclin-dependent kinase inhibitor p21 in human leukaemia HL-60 cells. The induction of p21 was associated with the accumulation of hypophosphorylated retinoblastoma protein (pRB) and the suppression of c-myc gene expression. Since the p53 gene is deleted in HL-60 cells, the anti-cancer PG is suggested to inhibit cancer cell growth by inducing p21 via a p53-independent pathway. Unlike HL-60 cells, cisplatin-resistant HL-60/R-CP cells were insensitive to Delta7-PGA1 methyl ester. While c-myc expression was transiently suppressed, neither G1 arrest nor hypophosphorylation of pRB was observed with the anti-cancer PG. Plasma membrane vesicles from HL-60/R-CP cells showed an enhanced level of GS-X pump (ATP-dependent glutathione S-conjugate export pump) activity towards the glutathione S-conjugate of Delta7-PGA1 methyl ester (Km 110 nM). GIF-0019 ¿N-carbomethoxy-S-[5-(4-benzoylphenyl)pentyl]glutathione dimethyl ester¿, a specific inhibitor of the GS-X pump, dose-dependently enhanced the cellular sensitivity of HL-60/R-CP cells to Delta7-PGA1 methyl ester and induced G1 arrest. The GS-X pump is suggested to play a pivotal role in modulating the biological action of the anti-cancer PG.

Inhibition of growth in oral squamous carcinoma cells by cyclopentenone prostaglandins: comparison with chemotherapeutic agents.

Four cyclopentenone prostaglandins (CPPGs) and PGE2 caused significant dose-dependent inhibition in growth of human oral squamous carcinoma cells (SCC-15). The rank order of their potency was PGJ2>PGA1>16, 16-dimethyl PGA1>PGA2>PGE2. In a follow-up experiment it was found that the mean per cent inhibition in cell growth by PGJ2 and delta12-PGJ2 at 10(-5) M was 61.22 and 63.81, while that of 5-fluorouracil and methotrexate was 36.67 and 38.86, respectively. delta12-PGJ2 and PGJ2 induced significant dose-dependent inhibition in nuclear DNA synthesis (i.e. cell proliferation). Combining vitamin E succinate with lower concentrations of CPPGs enhanced significantly their inhibitory effect on nuclear DNA synthesis of cancer cells.

Induction of MRP/GS-X pump and cellular resistance to anticancer prostaglandins.

We provide evidence that the expression of the human MRP/GS-X pump encoded by the MRP (multidrug resistance associated protein) gene is induced by cisplatin in human leukemia HL-60/R-CP (cisplatin-resistant) cells and modulates cell growth inhibition by delta(7)-prostaglandin A1 (PGA1) methyl ester. The MRP mRNA level in HL-60/R-CP cells increased remarkably after a 24-h incubation with 20 microM cisplatin; interestingly, however, no amplification of the MRP gene was detected. In cisplatin-sensitive HL-60 cells, which express the MRP/GS-X pump at low levels, c-myc expression was substantially suppressed by delta(7)-PGA1 methyl ester and the cell cycle was arrested in G1 phase. By contrast, in HL-60/R-CP cells overexpressing the MRP/GS-X pump, c-myc expression and cell proliferation were much less affected by delta(7)-PGA1 methyl ester. This suggests that induction of the MRP/GS-X pump may confer on cancer cells resistance to anticancer prostaglandins and that the resistance mechanism may involve the increased efflux of PG-glutathione conjugates, as active intermediates, from the cells via the MRP/GS-X pump.

Cell cycle arrest by prostaglandin A1 at the G1/S phase interface with up-regulation of oncogenes in S-49 cyc- cells.

Our previous studies have implied that prostaglandins inhibit cell growth independent of cAMP. Recent reports, however, have suggested that prostaglandin arrest of the cell cycle may be mediated through protein kinase A. In this report, in order to eliminate the role of c-AMP in prostaglandin mediated cell cycle arrest, we use the -49 lymphoma variant (cyc-) cells that lack adenylate cyclase activity. We demonstrate that dimethyl prostaglandin A1 (dmPGA1) inhibits DNA synthesis and cell growth in cyc- cells. DNA synthesis is inhibited 42% by dmPGA1 (50 microM) despite the fact that this cell line lacks cellular components needed for cAMP generation. The ability to decrease DNA synthesis depends upon the specific prostaglandin structure with the most effective form possessing the alpha, beta unsaturated ketone ring. Dimethyl PGA1 is most effective in inhibiting DNA synthesis in cyc- cells, with prostaglandins PGE1 and PGB1 being less potent inhibitors of DNA synthesis. DmPGE2 caused a significant stimulation of DNA synthesis. S-49 cyc- variant cells exposed to (30-50 microns) dmPGA1, arrested in the G1 phase of the cell cycle within 24 h. This growth arrest was reversed when the prostaglandin was removed from the cultured cells; growth resumed within hours showing that this treatment is not toxic. The S-49 cyc- cells were chosen not only for their lack of adenylate cyclase activity, but also because their cell cycle has been extensively studied and time requirements for G1, S, G2, and M phases are known. Within hours after prostaglandin removal the cells resume active DNA synthesis, and cell number doubles within 15 h suggesting rapid entry into S-phase DNA synthesis from the G1 cell cycle block.(ABSTRACT TRUNCATED AT 250 WORDS)

Effects of dimethyl prostaglandin A1 on herpes simplex virus and human immunodeficiency virus replication.

We have investigated the direct effect of dimethyl prostaglandin A1 (dmPGA1) on the replication of herpes simplex virus (HSV) and human immunodeficiency virus type 1 (HIV-1). dmPGA1 significantly inhibited viral replication in both HSV and HIV infection systems at concentrations of dmPGA1 that did not adversely alter cellular DNA synthesis. The 50% inhibitory concentration (ID50) for several HSV type 1 (HSV-1) strains ranged from 3.8 to 5.6 micrograms/ml for Vero cells and from 4.6 to 7.3 micrograms/ml for human foreskin fibroblasts. The ID50s for two HSV-2 strains varied from 3.8 to 4.5 micrograms/ml for Vero cells; the ID50 was 5.7 micrograms/ml for human foreskin fibroblasts. We found that closely related prostaglandins did not have the same effect on the replication of HSV; dmPGE2 and dmPGA2 caused up to a 60% increase in HSV replication compared with that in untreated virus-infected cells. HIV-1 replication in acutely infected T cells (VB line) and chronically infected macrophages was assessed by quantitative decreases in p24 concentration. The effective ID50s were 2.5 micrograms/ml for VB cells acutely infected with HIV-1 and 5.2 micrograms/m for chronically infected macrophages. dmPGA1 has an unusual broad-spectrum antiviral activity against both HSV and HIV-1 in vitro and offers a new class of potential therapeutic agents for in vivo use.

Selection of HTLV-I positive clones is prevented by prostaglandin A in infected cord blood cultures.

Type A prostaglandins (PGA1 and 16,16-dimethyl-PGA2-methyl ester) were found to block the proliferation of HTLV-I infected cord blood lymphocytes (CBL) in vitro, thus preventing the clonal immortalisation that is considered as a predisposing condition to HTLV-I positive leukaemia. PGA1 and di-M-PGA2 did not affect the long-term survival of normal non-infected CBL, whereas they suppressed the proliferation of an established cord-blood derived HTLV-I positive cell line, MT-2. As shown by the number of HTLV-I infected p19+ cells, the block of the selection of immortalised, infected clones by PGAs did not appear to be due to an inhibition of early stages of HTLV-I infection. The possibility that the effect of PGAs could be mediated by an action on the immune response was also examined. PGAs regulated the cell-mediated cytotoxic function of CBL to a different extent when normal non-infected or HTLV-I exposed CBL were compared. In fact, PGAs down-regulated the natural killing and macrophage/lymphocyte cytotoxic response of normal CBL, whereas they did not modify the already depressed immune response of CBL challenged with HTLV-I. These results suggest that the protective effect of PGAs against HTLV-I infection in vitro is mostly related to the direct suppression of the clonal expansion of virus-infected cells, rather than to the anti-viral activity or modulation of the cell-mediated immunity.

Involvement of prostaglandin-induced proteins in the inhibition of herpes simplex virus replication.

Cyclopentenone prostaglandin (PG), delta7-PGA1 was found to induce several polypeptides in human embryonic fibroblast (HEF) cells which were noticed to be dose-related and appeared after 1 h of treatment with a peak at around 5 h and gradual disappearance after 12 h. PG-induced proteins were almost identical in terms of molecular weights with those induced by heat-shock at 42 degrees C. Regarding the mechanism of inhibition of herpes simplex virus (HSV) replication by PG in cell culture, dot blot hybridization has revealed that the level of immediate early (IE) mRNA of the virus was reduced after PG treatment with time dependence. And this delayed inhibitory effect of delta7-PGA1 on HSV was shown to be associated with the production and accumulation of the induced polypeptides.

Polycyclic aromatic hydrocarbon quinones and glutathione thioethers as substrates and inhibitors of the human placental NADP-linked 15-hydroxyprostaglandin dehydrogenase.

The human placental NADP-linked 15-hydroxyprostaglandin dehydrogenase catalyzes oxidoreduction at the 9- and 15-positions of many prostaglandins, but its catalytic efficiency (i.e. kcat/Km) for these reactions is low (Jarabak, J., Luncsford, A., and Berkowitz, D. (1983) Prostaglandins 26, 849-868). In the present study, we demonstrate that both K-region and non-K-region o-quinones of polycyclic aromatic hydrocarbons are excellent substrates for this enzyme. These compounds are reduced with kcat/Km values ranging from 3 to 20 X 10(6) S-1 M-1. The glutathione thioethers of menadione and toluquinone are reduced with similar catalytic efficiencies. Furthermore, these substances and certain other glutathione thioethers are potent inhibitors of prostaglandin B1 oxidation ([I50] = 7 X 10(-8) to 5 X 10(-6) M); while several glutathione thioethers also inhibit polycyclic aromatic hydrocarbon quinone reduction ([I50] = 1.7-6.5 microM). These findings raise the possibility that the potential toxicity of quinones of polycyclic aromatic hyrocarbons and other xenobiotic substances may be altered in the placenta by an oxidoreductase for which prostaglandins are relatively poor substrates. They also suggest that the presence in placental tissue of certain glutathione thioethers could influence the reduction of these quinones and other xenobiotic substances by this enzyme.

Mechanism of inhibition of herpes simplex virus replication by delta 7-prostaglandin A1 and delta 12-prostaglandin J2.

We studied the effect of prostaglandins (PGs) A1, delta 7-A1, A2, D2, E1, E2, F2 alpha, J2 and delta 12-J2 on the replication of herpes simplex virus type 2 (HSV-2). Of nine PGs we tested, delta 7-PGA1 was found to have the most potent inhibitory effect; 50% inhibitory dose (ID50) was 0.35 microgram/ml in the plaque reduction assays and HSV-2 induced protein synthesis was strongly suppressed at 0.5 microgram/ml whereas at this dose, the protein synthesis of uninfected cells was not inhibited. Dot blot hybridization analysis revealed that delta 7-PGA1 and delta 12-PGJ2 inhibited the primary transcription of HSV-2. Thus we suggest that those PGs are primarily active at the level of mRNA synthesis.

Inhibition of the proliferation of transformed epidermal cells in culture by various prostaglandins.

Cytotoxic action of various prostaglandins (PGs) was examined on the PAM 212 transformed mouse epidermal cell line, and delta 7-PGA1 was found most active. delta 7-PGA1 exerted a dose-dependent inhibition of PAM 212 cell growth over 0.1 microgram/ml (0.3 microM). At 1.6 microgram/ml (4.6 microM) growth was completely inhibited, and the number of viable cells decreased remarkably during culture. The concentration needed for 50% growth inhibition (IC50) value of delta 7-PGA1 on PAM 212 cell growth was calculated as 0.4 microgram/ml (1.1 microM). At this concentration, the DNA synthesis in 24- and 48-h cultured cells was decreased to a half of the level in the control cells, and microscopically, remaining cells showed degenerative changes with many vacuoles in their cytoplasm. Prostaglandin D2, a major PG in mast cells, also showed potent cytotoxic activity. However, this action was expressed as 9-deoxy-delta 9,12-13,14-dihydro-PGD2 (delta 12-PGJ2), which was converted from PGD2 in plasma, and had a 3-fold stronger growth inhibitory activity than PGD2; the IC50 values of PGD2 and delta 12-PGJ2 were 2 micrograms/ml (5.7 microM) and 0.75 microgram/ml (2.1 microM), respectively. Among other PGs tested, PGA2 showed a comparable growth inhibitory activity, and PGB2, PGE1, and PGE2 less but significant activity. Prostaglandin F2 alpha and PGI2 however, had no such effect on cell proliferation at 5 micrograms/ml (14.3 microM) concentration, suggesting that cyclopentenone structure is an essential moiety of PG derivatives for cell growth inhibition. This cytotoxic action of delta 7-PGA1 and delta 12-PGJ2 appears to be independent of cyclic-AMP, since these PGs were virtually inactive in raising intracellular cyclic-AMP levels in PAM 212 cells.

Suppressive effects of various antitumor prostaglandin derivatives on the activity levels of DNA polymerases in human cultured tumor cells.

Some PG derivatives have been shown to be inhibitory for tumor cell growth. To elucidate the underlying mechanism(s) for this, we examined various DNA polymerase activities in PG-treated cultured cells. Human KBIII cells were exposed for one day to each of PGJ2, delta 7-PGA1 and delta 12-PGJ2 at a concentration of 5 micrograms/ml. The cells were harvested and homogenized, and the cell-free extracts were analyzed by phosphocellulose column chromatography for separation, identification, and quantification of each of the DNA polymerases. The results obtained were as follows: The activity of DNA polymerase alpha, which is responsible for DNA replication, has remained almost unchanged by any of the PG derivatives tested (77-87%), whereas the activity of DNA polymerase beta (repair enzyme) was found to dramatically decrease to 13 to 20% of that of the nontreated control cells by treatment with any of these PG derivatives. The results indicate that (a) these PG derivatives inhibit, either directly or indirectly, the synthesis of DNA polymerase beta in the KBIII cell, and (b) DNA polymerase beta may also participate in DNA replication other than DNA repair.

The effects of prostaglandins F2 alpha and E2 on the motility of the cat colon in vitro.

Controversy exists with respect to the motor effects of prostaglandins (PG) in the colon. We studied the actions of the prostaglandins F2 alpha and E2 on the spontaneous and stimulated contractions of the circular muscle of the cat colon in vitro. Both prostaglandins enhanced contractile activity. PG F2 alpha was more effective than PG E2, the ED50 were 4.2 X 10(-9) M and 2 X 10(-7) M, respectively. The prostaglandin antagonist 8 ethoxycarbonyl-5-methyl-10,11-dihydroxy-PG A1-ethyl-ester (HR 546) in a concentration of 10(-7) M shifted the ED50 of PG F2 alpha to the right by 1 log unit and unmasked motor stimulatory actions of low PG F2 alpha concentrations (10(-11) and 10(-10) M). The latter effect was atropine sensitive. In conclusion, PG F2 alpha and PG E2 stimulate contractions of the circular colon muscle at concentrations naturally occurring in the gut. It is conceivable that they play a role in the control of colonic motility, particularly in diseases like ulcerative colitis and Crohn's disease.