USOO9381193B2 (12) United States Patent (10) Patent No.: US 9,381,193 B2 Moley et al. (45) Date of Patent: Jul. 5, 2016 (54) CONTRACEPTIVE METHODS AND and during IVF treatment improves the pregnancy rate: A random COMPOSITIONS ized prospective study', 2010, vol. 25, No. 10, pp. 2496-2500.* WordNet Search 3.0, "prevent', also available at http://wordnetweb. princeton.edu/perl/webwn?s=prevent&sub=Search+WordNet&o2= (75) Inventors: Kelle H. Moley, St. Louis, MO (US); &o0=1&o8=1&o1=1&o7=&o5=&o9=&o6=&o3=&o4=&h=; last Antonina I. Frolova, St. Louis, MO viewed Dec. 10, 2013.* (US) Sander, Valeria et al., Neuroimmunomodulation, “The Influence of Dehydroepiandrosterone on Early Pregnancy in Mice”, 2005, vol. 12, (73) Assignee: WASHINGTON UNIVERSITY, St. pp. 285-292.* Schelbach, Cheryl J. et al., Reproduction, Fertility and Development, Louis, MO (US) “Mechanics contributing to the reduced developmental competence of glucosamine-exposed mouse oocytes', 2010, vol. 22, pp. 771 *) NotOt1Ce: Subjubject to anyy d1Sclaimer,disclai theh term off thisthi 779.3 patent is extended or adjusted under 35 Sourla, Antigone et al., J. Steroid Biochem. Molec. Biol. “Effect of U.S.C. 154(b) by 22 days. Dehydroepiandrosterone on Vaginal and Uterine Histomorphology in the Rat', 1998, vol. 66, No. 3, pp. 137-149.* (21) Appl. No.: 13/436,469 Suh-Burgmann, Elizabeth et al., Gynecol Obstet Invest, “Long-term Administration of Intravaginal Dehydroepiandrosterone on Regres sion of Low-Grade Cervical Dysplasia—A Pilot Study', 2003, vol. (22) Filed: Mar. 30, 2012 55, pp. 25-31.* Chamberlain, J. G. "Deleterious Effects of 6-Aminonicotinamide on (65) Prior Publication Data Implantation and Early Embryonic Development in Long-Evans US 2013/OO65853 A1 Mar. 14, 2013 Rat.” Anatomical Record, 1963, vol. 145. No. 2..., abstract. Elia, Evelin et al., International Immunopharmacology, “Detrimental Related U.S. Application Data effects of hyperandrogenism on utrine functions”, 2008, vol. 8, pp. 1827-1834. (60) Provisional application No. 61/470.925, filed on Apr. Frolova, Antonina et al., Endocrinology, "Facilitative Glucose Trans 1, 2011. porter Type 1 is Differentially Regulated by Progesterone and Estro gen in Murine and Human Endometrial Stromal Cells”, 2009, vol. 150, No. 3, pp. 1512-1520.* (51) Int. Cl. Hackl, Heinrich, Acta ObstetGynec Scand, "Metabolism of Glucose A6 IK3I/5685 (2006.01) in the Human Endometrium with Special Reference to Fertility and A6IP 5/8 (2006.01) Contraception”, 1973, vol. 52, pp. 135-140.* A6 IK3 L/455 (2006.01) Tuckerman, E. M. et al., Fertility and Sterility, “Do Androgens Have A6 IK3I/7008 (2006.01) a Direct Effect on Endometrial Function? An In Vitro Study', 2000, (52) U.S. Cl. vol. 74, No. 4, pp. 771-779.* CPC ........... A61 K3I/455 (2013.01); A61 K3I/5685 Ramathal, Cy, et al., “Endometrial Decidualization: Of Mice and Men'. Semin Reprod Med., Jan. 2010, 28(1): 17-26, p. 2. (2013.01); A61K3I/7008 (2013.01) Frolova, Dehydroepiandrosterone Inhibits Glucose FluxThrough the (58) Field of Classification Search Pentose Phosphate Pathway in Human and Mouse Endometrial CPC ... A61K 3 1/70; A61 K31/7008: A61K 31/45; Stromal Cells, Preventing Decidualization and Implantation, Mol A61K 3 1/566; A61K 31/08: A61K 3 1/655; Endocrinol, Aug. 2011, 1444-1455, 25 (8). A61K 31/166; A61 K31/70041: A61 K Kara, Does dehydroepiandrosterone Supplementation really affect 31/5685; A61K 31/455 IVF-ICSI outcome in women with poor ovarian reserve?, European USPC .............. 514/23, 62, 178,221, 352, 707, 179 Journal of Obstetrics & Gynecology and Reproductive Biology, See application file for complete search history. 2014, 63-65, 173. Narkwichean. Efficacy of dehydroepiandrosterone to improve ovarian response in women with diminished ovarian reserve: a meta (56) References Cited analysis, Reproductive Biology and Endocrinology, 2013, 8 pgs, U.S. PATENT DOCUMENTS 11:44. Tsi, Glucosamine inhibits decidualization of human endometrial 5.439,913 A * 8/1995 Chwalisz ............... 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Boca Raton, Florida. Guay, Testosterone insufficiency in women: factor fiction?, World J Urol, 2002, pp. 106-110, vol. 20. * cited by examiner U.S. Patent Jul. 5, 2016 Sheet 1 of 44 US 9,381,193 B2 1.4 - 1 - 0.8 - 0.6 - 0.4 - 0 --- ~f~ ~~ ~~ ---- ~~ ~ 0.5 1.0 2.5 50 O.O 17.5 Glucose concentration in media (m) FG 1A PRL higFBP O 0..48 6 0. 2 0.5 1. 2.5 5.0 10.0 17.5 Glucose concentration (my) F.G. 1B U.S. Patent Jul. 5, 2016 Sheet 2 of 44 US 9,381,193 B2 1.2 : O 0.5 1 15 lodoacetate concentration (m) F.G. 2A 1 4.68O2OOOOO 2O O O 0.5 1 1.5 lodoacetateIII concentration (mR) FG. 2B U.S. Patent Jul. 5, 2016 Sheet 3 of 44 US 9,381,193 B2 3. & & reast set is ratic ci&ys: a was a Control on Decidualized «» Decid+ 100. MDHEA FG 3A wa 3 3 extreet is statics (8x & 188 DeCicialized ** Decid+100MDHEA F.G. 3B U.S. Patent Jul. 5, 2016 Sheet 4 of 44 US 9,381,193 B2 FG. 3C Control Decidualized +100DHEA U.S. Patent Jul. 5, 2016 Sheet 5 of 44 US 9,381,193 B2 2 i O.8 O6 0,4 se 0.2 O. O Q W s' s°s S & S 8 { 2 s sN KS a C2 PR CFBP : 0.8 . U.S. Patent Jul. 5, 2016 Sheet 6 of 44 US 9,381,193 B2 8 3 8 rearrarit is ratic: iays a on an a Control mm Decidualized -o-o-o- Decid+10M 6AN FG 4A S. k 3 8 g s 5 o 4. e s 2 3 6 9 Treatment duration (days) as sees as Control -H Decidualized ass Decid+1 OuM 6AN F.G. 4B U.S. Patent Jul. 5, 2016 Sheet 7 of 44 US 9,381,193 B2 R IGFBP 0.8 e 0.6 : O4 O2 0.0 -r-----------s --- S ---- ^ -- CCS & NS' QS S U.S. Patent Jul. 5, 2016 Sheet 8 of 44 US 9,381,193 B2 <-- B-ACTIN Scrambled ShrNA G6PD shRNA - - - - MPA+db-CAMP - - - - i2 --- 8 FG 5B s Oa. 0.2 . Scrambled ShrNA t + As G6PD shrnA MPA+db-cAMP * Pr: nig?bpt FIG. 5C OO 68O 0. 4. .2 O Scrambled ShrNA t r G6PD ShrNA - - WPA-C-CAMP U.S. Patent Jul. 5, 2016 Sheet 9 of 44 US 9,381,193 B2 1-(4C)-Glucose 6-(14C)-Glucose 4. O 2.3. OO O 0. Contro Decidualized Decidualized Decidualized +2M 6-AN +5M 6-AN F.G. 6A 4.0 23 OO 0 0. U.S. Patent Jul. 5, 2016 Sheet 10 of 44 US 9,381,193 B2 250 200 1 5 0 O 5O U.S. Patent Jul. 5, 2016 Sheet 11 of 44 US 9,381,193 B2 Onset of o-> Oil Harvest implantation injection uterine horns DHEA-treated FG 7B U.S. Patent Jul. 5, 2016 Sheet 12 of 44 US 9,381,193 B2 O.25 Citro X.A. 0.20 3.35 .30 CO instiniated hors Oeciduofia FG 7C FG 7D U.S. Patent Jul. 5, 2016 Sheet 13 of 44 US 9,381,193 B2 Unst lated Dec iduoma FG 7E U.S. Patent Jul. 5, 2016 Sheet 14 of 44 US 9,381,193 B2 FG. 7F FG 7H U.S. Patent Jul. 5, 2016 Sheet 15 of 44 US 9,381,193 B2 DHEA F.G. 8A U.S. Patent Jul. 5, 2016 Sheet 16 of 44 US 9,381,193 B2 0.2 : sk Contro DHEA 0.15 G :s 0.1 : s 0.05 0 ico. --------------------- ----------- Control Deciduonna F.G. BB ck 0.25 - Control U.S. Patent Jul. 5, 2016 Sheet 17 of 44 US 9,381,193 B2 Control 100 MDHEA 50M 6AN FG 9A Control 50MDHEA - XX X X X X XX 100M DHEA FG 9B U.S.