Novel contraceptive targets to inhibit ovulation: the prostaglandin E2 pathway

Abstract

Background: Prostaglandin E2 (PGE2) is an essential intrafollicular regulator of ovulation. In contrast with the one-gene, one-protein concept for synthesis of peptide signaling molecules, production and metabolism of bioactive PGE2 requires controlled expression of many proteins, correct subcellular localization of enzymes, coordinated PGE2 synthesis and metabolism, and prostaglandin transport in and out of cells to facilitate PGE2 action and degradation. Elevated intrafollicular PGE2 is required for successful ovulation, so disruption of PGE2 synthesis, metabolism or transport may yield effective contraceptive strategies.

Methods: This review summarizes case reports and studies on ovulation inhibition in women and macaques treated with cyclooxygenase inhibitors published from 1987 to 2014. These findings are discussed in the context of studies describing levels of mRNA, protein, and activity of prostaglandin synthesis and metabolic enzymes as well as prostaglandin transporters in ovarian cells.

Results: The ovulatory surge of LH regulates the expression of each component of the PGE2 synthesis-metabolism-transport pathway within the ovulatory follicle. Data from primary ovarian cells and cancer cell lines suggest that enzymes and transporters can cooperate to optimize bioactive PGE2 levels. Elevated intrafollicular PGE2 mediates key ovulatory events including cumulus expansion, follicle rupture and oocyte release. Inhibitors of the prostaglandin-endoperoxide synthase 2 (PTGS2) enzyme (also known as cyclooxygenase-2 or COX2) reduce ovulation rates in women. Studies in macaques show that PTGS2 inhibitors can reduce the rates of cumulus expansion, oocyte release, follicle rupture, oocyte nuclear maturation and fertilization. A PTGS2 inhibitor reduced pregnancy rates in breeding macaques when administered to simulate emergency contraception. However, PTGS2 inhibition did not prevent pregnancy in monkeys when administered to simulate monthly contraceptive use.

Conclusion: PTGS2 inhibitors alone may be suitable for use as emergency contraceptives. However, drugs of this class are unlikely to be effective as monthly contraceptives. Inhibitors of additional PGE2 synthesis enzymes or modulation of PGE2 metabolism or transport also hold potential for reducing follicular PGE2 and preventing ovulation. Approaches which target multiple components of the PGE2 synthesis-metabolism-transport pathway may be required to effectively block ovulation and lead to the development of novel contraceptive options for women. Therapies which target PGE2 may also impact disorders of the uterus and could also have benefits for women's health in addition to contraception.

Keywords: contraception; follicle; ovary; prostaglandin E2, cyclooxygenase-2.

Figure 1

Structural changes in the dominant follicle during ovulation. Before the LH surge, the oocyte (pink) is surrounded by the zona pellucida (gray) and tight cumulus granulosa cells (light green); mural granulosa cells (dark green) surround the follicular fluid (yellow). Outside the granulosa cell basement membrane (gray), stromal components include theca cells (purple) and small vessels (red). The follicle apex is covered by the ovarian surface epithelium (blue). In response to the LH surge, the oocyte resumes meiosis and produces the first polar body. The cumulus expands and detaches from the mural granulosa cells. Luteinization includes hypertrophy of granulosa cells and invasion of theca cells and new vessels into the granulosa cell layer. A breach in the ovarian surface epithelium, underlying stroma, and mural granulosa cells results in rupture at the follicle apex. Mural granulosa cells protrude through the rupture site to form an ovulatory stigmata. The ovulatory canal permits release of the cumulus-oocyte complex (COC).

Figure 2

Enzymes involved in PGE2 synthesis and metabolism. Arachidonic acid is cleaved from membrane phospholipids by a phospholipase A2 (PLA2). A cyclooxygenase (PTGS1 or PTGS2) converts arachidonic acid to prostaglandin H2 (PGH2). PGH2 can be converted to PGE2 by a PGE2 synthase (PTGES) or to PGF2α by aldo-keto reductase 1C3 (AKR1C3). PGE2 is metabolized to PGF2α by an enzyme with 9-keto reductase activity (AKR1C1 or AKR1C2). PGE2 is metabolized to 15-keto-PGE2 by prostaglandin dehydrogenase (HPGD). For each conversion, enzyme listed in boldface is the predominant form in the ovulatory follicle. Molecular structures are provided to highlight the conversion made by each enzyme. PGE1, PGE2 and PGE3 are distinguished by the number of double bonds in the molecular structure.

Figure 3

PGE2 synthesis and metabolism enzyme mRNA, protein, and activity in primate ovulatory follicles. All cells and tissues were obtained after ovarian stimulation before (0 h) and 12, 24, or 36 h after administration of an ovulatory dose of hCG. Ovulation is anticipated 37–42 h after the ovulatory gonadotrophin stimulus, so cell/tissue collections at times 0–36 h after hCG span the ovulatory period. Levels of mRNA (A–D), protein (E–H) and activity (I and J) for PLA2G4A (A, E and I), PTGS2 (B and F), PTGES (E and G), and 15-hydroxyprostaglandin dehydrogenase (HPGD) (D, H and J) in monkey granulosa cells are shown. PGE2 (K) and 15-keto PGE2 metabolites (PGEM; L) levels in follicular fluid from monkey ovulatory follicles are shown. (M) Relative levels of PGE2 synthesis and PGE2 metabolism enzyme activities indicate that metabolism predominates early in the ovulatory period while synthesis predominates late in the ovulatory period. In (A)–(D), (G), and (I–L), groups with no common letters (a,b,c) are different by analysis of variance and appropriate post hoc analysis. In (E) and (F), an = antrum, gc = granulosa cell, st = stroma, asterisk indicates luteinizing granulosa cell layer, arrowhead indicates stromal (possibly theca) cell immunopositive for PTGS2. In (I), nd = not determined. See text for details; republished with permissions from Duffy and Stouffer (2001), Duffy et al. (2005a, b, c) and Duffy (2011).

Figure 4

PGE2 synthesis, action, transport and metabolism in granulosa cells. PGE2 is synthesized by the enzymes PLA2G4A, PTGS2, and PTGES which are associated with intracellular membranes including the endoplasmic reticulum (ER). Intermediates in PGE2 synthesis include arachidonic acid (AA) and PGH2. Functional PGE2 receptors (PTGERs) can be located in the plasma membrane or nuclear envelope, so both intracellular and extracellular PGE2 can bind to receptors and initiate signal transduction. PGE2 can exit and enter a cell via passive diffusion through the plasma membrane or facilitated diffusion via transporters such as SLCO2A1 and MRP4. Bioactive PGE2 is converted to the inactive PGE metabolite 15-keto-PGE2 through the activity of the cytoplasmic enzyme HPGD.

Figure 5

Ultrasound evaluation of ovulatory failure in women. An ovarian follicle in a woman receiving placebo was at 19 mm diameter on the day of the LH peak (A) and showed decreased follicular diameter (follicle collapse; consistent with follicle rupture) on the day after peak LH (B). An ovarian follicle in a woman receiving the PTGS2 inhibitor rofecoxib was at 22 mm diameter on the day of peak LH (C) and was at 50 mm diameter on Day 10 after peak LH (D). With PTGS2 inhibitor treatment, the follicle continued to increase in diameter after the LH surge (no follicle collapse; failure of follicle rupture). In these ultrasound images, ovarian follicles are black circles, with dotted lines to indicate measurement of follicular diameters. All panels are at approximately the same magnification. Republished with permissions from Pall et al. (2001).