Prostaglandin A1 Decreases the Phosphorylation of Tau by Activating Protein Phosphatase 2A via a Michael Addition Mechanism at Cysteine 377.

Prostaglandin (PG) A1 is a metabolic product of cyclooxygenase 2 (COX-2) that is potentially involved in regulating the development and progression of Alzheimer's disease (AD). PGA1 is a cyclopentenone (cy) PG characterized by the presence of a chemically reactive α,ß-unsaturated carbonyl. PGA1 is potentially involved in the regulation of multiple biological processes via Michael addition; however, the specific roles of PGA1 in AD remain unclear. TauP301S transgenic (Tg) mice were used as in vivo AD models, and neuroblastoma (N) 2a cells were used as an in vitro neuronal model. The PGA1-binding proteins were identified by HPLC-MS-MS after intracerebroventricular injection (i.c.v) of PGA1. Western blotting was used to determine tau phosphorylation in PGA1-treated Tg mice in the absence or in the presence of okadaic acid (OA), an inhibitor of protein phosphatase (PP) 2A. A combination of pull-down assay, immunoprecipitation, western blotting, and HPLC-MS-MS was used to determine that the PP2A scaffold subunit A alpha (PPP2R1A) is activated by the direct binding of PGA1 to cysteine 377. The effect of inhibiting tau hyperphosphorylation was tested in the Morris maze to determine the inhibitory effects of PGA1 on cognitive decline in tauP301S Tg mice. Incubation with N2a cells, pull-down assay, and mass spectrometry (MS) analysis revealed and indicated that PGA1 binds to more than 1000 proteins; some of these proteins are associated with AD and especially with tauopathies. Moreover, short-term administration of PGA1 in tauP301S Tg mice significantly decreased tau phosphorylation at Thr181, Ser202, and Ser404 in a dose-dependent manner. This effect was caused by the activation of PPP2R1A in tauP301S Tg mice. Importantly, PGA1 can form a Michael adduct with cysteine 377 of PPP2R1A, which is critical for the enzymatic activity of PP2A. Long-term treatment of tauP301S Tg mice with PGA1 activated PP2A and significantly reduced tau phosphorylation resulting in improvements in cognitive decline in tauP301S Tg mice. Our data provided new insight into the mechanisms of the ameliorating effects of PGA1 on cognitive decline in tauP301S Tg mice by activating PP2A via a mechanism involving the formation of a Michael adduct with cysteine 377 of PPP2R1A.

Medical methods for mid-trimester termination of pregnancy.

BACKGROUND: With the improvement of ultrasound technology, the likelihood of detection of major fetal structural anomalies in mid-pregnancy has increased considerably. Upon the detection of serious anomalies, women typically are offered the option of pregnancy termination. Additionally, there are still many reasons other than fetal anomalies why women seek abortion in the mid-trimester. OBJECTIVES: To compare different methods of second trimester medical termination of pregnancy for their efficacy and side-effects. SEARCH STRATEGY: We searched the Cochrane Central Register of Controlled Trials (CENTRAL) (The Cochrane Library), MEDLINE, Popline and reference lists of retrieved papers and other sources. SELECTION CRITERIA: All randomised controlled trials (RCTs) examining medical regimens for termination of pregnancy of a singleton living fetus between 12-28 weeks' gestation were analysed. The outcome measures were the induction to abortion interval, abortion rate within 24 hours, need for surgical evacuation, blood loss, uterine rupture, pain, and side-effects.Trials including >20% fetal death, multiple pregnancies, previous uterine scars and regimens which involved cervical preparation were excluded. DATA COLLECTION AND ANALYSIS: Two authors selected the trials and three authors extracted data. MAIN RESULTS: Fourty RCTs were included, addressing various agents for pregnancy termination and methods of administration. When used alone, misoprostol was an effective inductive agent, though it appeared to be more effective in combination with mifepristone. However, the evidence from RCTs is limited.Misoprostol was preferably administered vaginally, although among multiparous women sublingual administration appeared equally effective. A range of doses of vaginally administered misoprostol has been used. No randomised trials comparing doses of misoprostol were identified; however low doses of misoprostol appear to be associated with fewer side-effects while moderate doses appear to be more efficient in completing abortion. Four RCTs showed that the induction to abortion interval with 3-hourly vaginal administration of prostaglandins is shorter than 6-hourly administration without an increase in side-effects.Many studies reported the need for surgical evacuation. Indications for surgical evacuation include retained products of the placenta and heavy vaginal bleeding. Fewer women required surgical evacuation when misoprostol was administrated vaginally compared with women receiving intra-amniotical PGF(2a) . Mild, self-limiting diarrhoea was more common among women who received misoprostol compared to other agents. AUTHORS' CONCLUSIONS: Medical abortion in the second trimester using the combination of mifepristone and misoprostol appeared to have the highest efficacy and shortest abortion time interval. Where mifepristone is not available, misoprostol alone is a reasonable alternative. The optimal route for administering misoprostol is vaginally, preferably using tablets at 3-hourly intervals. Apart from pain, the side-effects of vaginal misoprostol are usually mild and self limiting. Conclusions from this review are limited by the gestational age ranges and variable medical regimens, including dosing, administrative routes and intervals of medication, of the included trials.

Neuroprotective effects of prostaglandin A1 in animal models of focal ischemia.

The present study evaluated the neuroprotective potential of prostaglandin A1 (PGA1) in rodent models of focal cerebral ischemia. PGA1 33 nmol reduced infarction volume by about 43% (P < 0.05) when administered intracerebroventricularly before and after transient ischemia in mice. PGA1 16.5-66 nmol diminished infarction volume by 18% to 27% (P < 0.01) when administered immediately following permanent ischemia in rats. PGA1 treatment also significantly ameliorated motor dysfunction after brain ischemia. These results suggest that PGA1 protects neurons from ischemic injury.

A chromatographic method for the quantification of prostaglandin E(1) and prostaglandin A(1) encapsulated in an intravenous lipid formulation.

Peripheral vascular disease is a common ailment of the aged and diabetic communities. As the numbers of these individuals increase, the need for therapeutic interventions will continue to grow. One of the possible therapies is the use of prostaglandins (PGE(1), prostacyclin and Iloprost) to decrease the vascular tone and increase vascular blood flow. Due to the hydrophobicity of the prostaglandins and prostaglandin analogues, various vehicles have been utilized to maintain the active pharmaceutical ingredient in a stable solution, e.g. alpha-cyclodextrin (Alprostadil, Edex) or emulsified lipid vehicles. In our laboratory, we designed a method for separating and assaying lipid-encapsulated PGE(1). Utilizing organic extraction, automated solid-phase extraction and precipitation techniques, we validated the measurement of the PGE(1) and PGA(1) content of the clinical drug formulation in the microgram per milliliter concentration range with an high performance liquid chromatography (HPLC) assay.

Antitumor activity, optimum administration method and pharmacokinetics of 13,14-dihydro-15-deoxy-deoxy-Delta7 -prostaglandin A1 methyl ester (TEI-9826) integrated in lipid microspheres (Lipo TEI-9826).

13,14-Dihydro-15-deoxy-Delta7-prostaglandin A1 methyl ester (TEI-9826), an antitumor prostaglandin analog, is a candidate for clinical trial. In the present study, we examined its biological stability in vitro, antitumor activity in vitro and in vivo, and pharmacokinetics. Although TEI-9826 was rapidly hydrolyzed to the carboxylic acid form (TOK-4528), TOK-4528 as well as Delta12-prostaglandin J2 (PGJ2) were found to be stable in rat, mouse and human serum in vitro. TEI-9826 exhibited nearly identical or greater potential antitumor activity compared to Delta12-PGJ2 and Delta7-PGA1 in vitro against Colon26 tumor cells. Further evaluation of TEI-9826 using the 38 human cancer cell lines panel and COMPARE analysis suggested that its mode of action is quite different from other anticancer agents that are currently used. TEI-9826 was integrated into lipid microspheres (Lipo TEI-9826) for dosing. Growth inhibition by Lipo TEI-9826 against Colon26 tumor inoculated s.c. in mice depended on administration route, i.e. at 80 mg/kg, no growth suppressive effect was observed for daily bolus i.v., but significant growth suppressive effect was observed for daily i.p., daily s.c. every other day s.c. and 4 times a day continuous (5 min) i.v. These tumor growth-suppressive effects were cytostatic and the tumor started to regrow at the end or a few days after the end of administration. The pharmacokinetic study suggested that maintaining the blood level of TEI-9826 and/or TOK-4528 was essential for their antitumor effects. These results show that continuous i.v. infusion might be the most suitable administration method of Lipo TEI-9826 for clinical trial.

Preparation and evaluation of o/w type emulsions containing antitumor prostaglandin.

Antitumor prostaglandins(PGs) such as Delta12-PGJ2 and Delta7-PGA1 possess a cyclopentenone or cross-conjugated dienone structures. Antitumor PGs are actively incorporated through cell membrane and control gene expression. Very recent studies clarified that P53 independent expression of p21 and gadd 45, activation of PPARgamma are involved in antitumor mechanism of these PGs. At the low concentration, these PGs exhibit physiological or pathological activity such as osteoblast calcification, promotion of colon cancer cell proliferation. COMPARE PROGRAM using human 38 tumor cell lines suggested that antitumor mechanism of Delta7-PGA1 and 13, 14-dihydro-15-deoxy-Delta7-PGA1 methyl ester (TEI-9826) are quite different from other anticancer agents which are clinically used. Lipid microspheres and Lipiodol formulation were examined as dosage form of the PGs and lipid microspheres were selected for further study. At first lipid microspheres integrated TEI-9038 (Lipo TEI-9038) was chosen as a candidate for clinical trial. However Lipo TEI-9038 failed to exhibit substantial antitumor effect because of its enzymatic instability and toxicity in vivo. Lipo TEI-9826 was then selected as promising candidate for clinical trial because of its stability in serum. Lipo TEI-9826 exhibited marked antitumor effect in several animal models including CDDP resistant nude mice model. Pharmacokinetic and toxicological studies using rats suggested that continuous infusion is the most suitable administration method for Lipo TEI-9826. New type emulsifier, Controlled High Pressure Process Homogenizer (De-BEE 2000 and mini De-BEE) was developed during the preclinical studies on manufacturing process of Lipo TEI-9826. These results warrant the clinical trial for Lipo TEI-9826 in CDDP resistant cancer.

Antitumor activity of 13,14-dihydro-15-deoxy-delta7-prostaglandin-A1-methyl ester integrated into lipid microspheres against human ovarian carcinoma cells resistant to cisplatin in vivo.

One of the delta7-prostaglandin A1 derivatives with unique antitumor activities, 13,14-dihydro-15-deoxy-delta7-prostaglandin-A1-methyl ester, was integrated into lipid microspheres (Lipo-TEI9826) and examined for its antitumor effect in vitro and in vivo. The in vitro relative resistance of human ovarian cancer, A2780CP, to cisplatin (CDDP) and Lipo-TEI9826 was 27.3 and 2.0, respectively, compared with A2780, the parent cell line of A2780CP. In in vivo experiments, when A2780CP and the parent cell line A2780 were inoculated into nude mice, A2780CP grew two times more rapidly than did A2780. The growth of A2780CP tumor was not suppressed by CDDP, whereas that of the A2780 tumor was significantly suppressed. Nevertheless, the growth of both the A2780 and the A2780CP inoculated tumors was significantly inhibited by treatment with Lipo-TEI9826 at any time after the initial treatment, compared with the lipid microspheres only. These results show that Lipo-TEI9826 may be an effective antitumor agent and capable of overcoming CDDP resistance.

Effect of prostaglandin A1 on proliferation and telomerase activity of human melanoma cells in vitro.

Previous studies have shown that cyclopentenone prostaglandins are endowed with antitumour activity in various murine and human tumour models. In the present investigation four human melanoma cell lines were treated with graded concentrations (4-16microg/ml) of prostaglandin A1 (PGA1) for 24 or 48 h in vitro. At the end of the treatment, cell proliferation (measured in terms of DNA synthesis) and telomerase activity were determined. The results showed that PGA1 induced concentration-dependent inhibition of DNA synthesis at 48 h but not at 24 h in SK-MEL-28 cells. In contrast, marked inhibition of telomerase activity was detected after only 24 h of PGA1 treatment. Moreover, after 48h of treatment with the agent, inhibition of telomerase was more pronounced than inhibition of cell proliferation. Additional studies performed with three freshly generated melanoma cell lines confirmed that PGA1 produced early inhibition of cell growth accompanied by marked impairment of telomerase activity. These results suggest that PGA1 could be of potential value as antitumour agent, on the basis of two distinct mechanisms: direct cytostatic/cytotoxic effects on melanoma cells, and inhibitory activity on a tumour-associated enzymatic function (i.e. telomerase) that is responsible for cancer cell immortality.

Combined treatments with interferon (alpha,beta) plus PGA1 to control early infection with HTLV-I in primary cord blood-derived mononuclear cells.

Interferon (IFN) alpha and beta can activate an antiviral and immunomodulating response in primary cord blood-derived mononuclear cells (CBMC) exposed to infection with Human T-cell Leukemia Virus type I (HTLV-I), resulting in partial inhibition of early infection in vitro. On the other hand, PGA1, a PGE1-derived cyclopentenone prostaglandin, can inhibit in vitro the proliferation of virus-infected CBMC, preventing the emergence of the potentially transformed clone. In order to achieve a complete control of HTLV-I infection in this experimental model, we evaluated whether the antiviral activity of IFNs and the antiproliferative activity of PGA1 could be preserved in a combination therapy scheme. Recipient CBMC were treated with IFN alpha or beta (1000 IU/ml) at the onset of the co-culture with lethally irradiated virus-donor MT-2 cells, followed by multiple treatments with PGA1 (4 micrograms/ml every 4 days, starting on day 0) for 6 weeks post infection (p.i.). In PGA1-treated co-cultures the percentage of virus-positive CBMC was constantly doubled during culture time as well as the amount of viral transcripts and p19 virus core protein production were increased. The antiviral effects of IFNs, resulting in about a 50% reduction of the percentage of virus-positive CBMC and consequently in a partial inhibition of virus expression (HTLV-I transcription and p19 production) until 4 weeks p.i., were suppressed by multiple PGA1 treatments. However, the antiproliferative effect of PGA1 was enforced in IFN-treated co-cultures, leading to earlier control of proliferation of virus-infected cells. Interestingly, infection of CBMC with HTLV-I was associated with persistent expression of 70 kDa heat shock protein (HSP70), for at least 4 weeks p.i. IFNs and PGA1 showed antagonistic effects on HSP70 production in infected CBMC. In fact, production of HSP70 was suppressed (or prevented) in IFN-treated co-cultures, tested 2 and 4 weeks p.i. The fact that the expression of HSP70 is apparently suppressed (or prevented) by IFN treatment is surprising, since expression of this protein family has been associated with antiviral immunity. PGA1 could totally reverse the IFN-mediated suppression of HSP70 expression in these co-cultures. It is presently unclear whether HSP70 expression is directly involved in the control of proliferation exerted by PGA1 against virus-infected CBMC or is an epiphenomenon associated with inhibition of cell growth.

Effect of topical prostaglandin PGA2, PGA2 isopropyl ester, and PGF2 alpha isopropyl ester on intraocular pressure in normotensive and glaucomatous canine eyes.

Topical instillations of 1.0, 10, and 20 micrograms/50 microliters of prostaglandin PGA2, 0.5 and 1.0 microgram/50 microliters of PGA2 isopropyl ester, and 0.5, 1.0, 5.0 and 10.0 micrograms/50 microliters of PGF2 alpha isopropyl ester were evaluated in the normal dogs and glaucomatous beagles eyes. Each concentration of drug was evaluated for a seven day period. On Day 1 baseline values were obtained, days 2-4, the drug was instilled (once a day) and on days 5-7 post-treatment values were measured. All concentrations of PGA2 failed to lower intraocular pressure (IOP) in the normal and the glaucomatous (P greater than 0.72) dogs. PGA2 isopropyl ester decreased IOP in the normal dogs and in the glaucomatous beagles (P less than 0.01). The declines in IOP were significant at 1/2 to 1 hour and continued for up to 5 hours. No significant change in IOP occurred in the non-treated fellow eye of the normotensive dog (P less than 0.54) and the glaucomatous beagle (P less than 0.29). All concentrations of PGF2 alpha isopropyl ester significantly decreased IOP in the treated eyes of the normotensive dog (P less than 0.05) and the glaucomatous beagle (P less than 0.01). The significant change in IOP occurred within one hour after the instillation of PGF2 alpha isopropyl ester. The IOP remained lower than the baseline pressures 24 hours post-treatment for both the normotensive and glaucomatous dogs. Maximal change in IOP for normal dogs was a decrease of 9 mm Hg while the glaucomatous beagle had a decrease of 19 mm Hg. No significant change in IOP occurred in the non-treated fellow eye of the normotensive animal (P less than 0.16) and the glaucomatous beagle (P less than 0.40). The side effects of PGF2 alpha isopropyl ester were miosis and mild conjunctival irritation.

Augmentation of antiproliferative activity of recombinant human tumor necrosis factor by delta 12-prostaglandin J2.

The effects of tumor necrosis factor (TNF) and cyclopentenone prostaglandins (PGA2 and delta 12-PGJ2), singly and in combination, against proliferation in three human gynecologic tumor cell lines (HeLa S3, HHUA, and CAOV-3) were examined in vitro. The HeLa S3 and CAOV-3 cells were unresponsive to TNF, and the HHUA cells exhibited a minimal degree of responsiveness to TNF. The HeLa S3 and HHUA cells were responsive dose-dependently to PGA2, and the CAOV-3 cells were slightly responsive to PGA2. All three cell lines were highly responsive to delta 12-PGJ2. The synergistic antiproliferative effect of TNF and delta 12-PGJ2 appeared in all three cell lines. Pretreatment with delta 12-PGJ2 enhanced the effectiveness of TNF in these cell lines, but not vice versa. A synergistic interaction between TNF and PGA2 was absent in these three cell lines. These results suggest that a combined treatment with TNF and delta 12-PGJ2 could provide a new approach to obtaining increased responses in clinical trials.

Prolonged survival time of sarcoma 180-bearing mice treated with lipid microspheres-entrapped antitumor marine coral prostanoids.

Antitumor marine coral prostanoids (clavulone II and chlorovulone I) were entrapped into lipid microspheres of 0.2 micron diameter to make lipo-drugs. Daily treatment with lipo-chlorovulone I (1.6 mg/kg/day, i.p.) and lipo-clavulone II (12.5 mg/kg/day, i.p.) on days 1 through 5 markedly prolonged the survival time (135% ILS and 73% ILS, respectively) of mice inoculated with sarcoma 180 as compared with that of a corresponding dose of respective free chlorovulone I and clavulone II. These results suggest that lipid microspheres may be used as drug delivery carriers for antitumor coral prostanoids in vivo.

Eicosanoids as a new class of ocular hypotensive agents. 1. The apparent therapeutic advantages of derived prostaglandins of the A and B type as compared with primary prostaglandins of the E, F and D type.

The classic primary prostaglandins (PGs), as well as some of their analogs and derivatives, are potent ocular hypotensive agents. The present studies show that A and B PGs, which are derived from PGs of the E type by dehydration and isomerization, have a much greater ocular hypotensive potency than the primary PGs of the E, F or D type. A single application of 5 micrograms of PGA2 to the cat eye in a 25-microliters volume of aqueous vehicle solution yielded a greater and more prolonged ocular hypotensive effect than as much as 100 micrograms of topically applied PGF2 alpha. As little as 1 microgram of PGA2 had a significant ocular hypotensive effect that was enhanced by three or more consecutive daily applications of the same dose. This IOP reduction, which remained significant for several days after the last of 10 daily treatments, was not associated with biomicroscopically detectable flare or invasion of the anterior chamber by cells. Although PGF2 alpha and, to a much lesser extent, PGE2 have a miotic effect in cats, PGs of the A and B type did not cause significant miosis even at doses 50- to 100-fold greater than the minimum dose required to yield significant ocular hypotension. PGA2 retained its ocular hypotensive potency when stored in an aqueous solution at room temperature for four months. The conjunctival hyperemia caused by 5 micrograms or 10 micrograms of A or B type PGs on rabbit eyes was milder and shorter in duration than that caused by the same doses of PGE2 or PGF2 alpha. These findings suggest that derived PGs, especially PGs of the A type, may have a therapeutic advantage over primary PGs for the treatment of ocular hypertension and glaucoma.

A comparison of the effect of Lipo PGE1 and other related compounds on experimental peripheral arterial occlusive disease in rats.

The effect of Lipo PGE1 (prostaglandin E1 incorporated in lipid microspheres) on laurate-induced peripheral arterial occlusive disease in rats was evaluated and compared with that of other related compounds. Lipo PGE1 showed a potent inhibitory effect on the progression of lesions in this model, but Lipo PGA1 had no such effect. Moreover, the effect of Lipo PGE1 was more potent than that of PGE1 X CD (cyclodextrin clathrated PGE1) and PGE1 on this model. These results suggest that incorporation of PGE1 into soybean oil microspheres might exert a more potent effect by facilitating drug delivery to the lesions in this model. It would appear that Lipo PGE1 is suitable for clinical use in peripheral vascular disorders.

Dexamethasone, prostaglandin A, and retinoic acid modulation of murine and human melanoma cells grown in soft agar.

The cloning efficiencies of a murine melanoma cell line (S91 CCL 53.1) and a human melanoma cell strain (C8146c) were inhibited by dexamethasone (DEX), prostaglandin A1 (PGA1), and beta-all-trans-retinoic acid (RA) in a dose-dependent manner. Murine melanoma tumor colony-forming units (MTCFU) were inhibited more than 99% by DEX (1 X 10(-7) M) and RA (1 X 10(-7) M) with a concentration needed to produce a 50% reduction in colony formation for both hormones of 5 X 10(-9) M. Combinations of DEX and RA effected a synergistic inhibition on colony formation, which was reflected by a 11/2 log reduction in the hormone concentration needed to produce a greater than 99% inhibition of colony formation. When PGA1 was added to DEX and RA, a greater than additive reduction in colony formation was observed. Human MTCFU from cell strain C8146c were inhibited more than 85% at an RA concentration of 1 X 10(-7) M, but they were reduced only to 40% of control at a DEX concentration of 1 X 10(-6) M. DEX-RA produced an additive inhibition of colony formation. Addition of submaximal amounts of PGA1 to DEX-RA combinations or to either hormone alone resulted in synergistic reduction of human MTCFU. These results demonstrated that the proliferative potential of human and murine melanomas can be simultaneously regulated by DEX, PGA1, and RA.

Therapeutic use of PGA1 infusions in severe pre-eclampsia - a major clinical potential.

Pilot studies showed that, i.v. infusions of the renal prostaglandin A1 (PGA1) induced a triad of beneficial clinical responses in severe pre-eclampsia; the blood pressure became normotensive, renal function was markedly improved and labour was successfully induced. The present study was an attempt to develop a therapeutic schedule of PGA1 administration in severe toxemia. Twenty one cases of severe pre-eclampsia (in 3 equal groups) received i.v. infusions of PGA1 in a dose range of 0.1-0.5 microgram/kgm/min for 12 - 24 hours and the B.P., uterine activity and FHR were continuously monitored during and for 12 hours following the infusion period. The 0.1 microgram/Kgm/min dose for 12 hours was inadequate while 0.5 microgram/Kgm/min for 12 hours induced a good hypotensive response and the cases delivered within 48 hours but a post-infusion rebound in hypertension was observed. The dose of 0.5 microgram/Kgm/min for 24 hours appeared to be optimal in clinical terms since a satisfactory effect on B.P. was recorded and all the subjects delivered normal babies during the infusion period with minimal or no post-infusion rebound rise in B.P. This approach holds a major potential in the treatment of severe pre-eclampsia.

Hypertensive emergencies: practical approach to treatment.

In a hypertensive crisis, an antihyprtensive agent must be able to reduce blood pressure to safe levels rapidly and predictably. Parenteral therapy is usually necessary, and in general, agents that can be infused and titrated are preferable to those that cannot. This article tells which drugs to use in specific situations and gives explicit instructions for intravenous and oral administration.