Compounds for tocolytic use

Abstract

Disclosed herein are methods for treating or reducing symptoms of preterm labor or decreasing preterm birth in a subject. The methods include administering to the subject a therapeutically effective amount of a compound for tocolytic use.

Description

VU Ref.24027 / MBF Ref.093386-0031-WO01 COMPOUNDS FOR TOCOLYTIC USE CROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application claims priority to U.S. Provisional Patent Application No.63 / 591,085 filed October 17, 2023 and U.S. Provisional Patent Application No.63 / 591,423 filed October 18, 2023, each of which is incorporated herein by reference in its entirety. STATEMENT REGARDING FEDERALLY SPONSORED RESEARCH

[0002] This invention was made with government support under grant HD098213 awarded by the National Institutes of Health. The government has certain rights in the invention. FIELD

[0003] This disclosure relates to drugs for tocolytic use and methods for the prevention or treatment for preterm labor. INTRODUCTION

[0004] Preterm birth (PTB), resulting from preterm labor, is a significant global health problem affecting 5-18% of pregnancies worldwide and is associated with 60-80% of infant mortality and 50% neonatal morbidities. Currently, there is a lack of FDA-approved tocolytics for the management of preterm labor. Drugs (nifedipine, terbutaline, and indomethacin) used off-label for tocolytic use to suppress labor were initially developed to treat disorders in other smooth muscle types such as vascular and pulmonary. Thus, the cardiovascular side-effects (maternal: myocardial ischemia, hypertension / hypotension, arrhythmias and tachycardias^ fetal: in utero constriction of the ductus arteriosus (DA) and tachycardia) if used greater than 48-72 hours are not surprising. Uterospecific drug development has yielded: (i) oxytocin-receptor (OXTR) antagonists, such as atosiban, for tocolytic use and (ii) a synthetic progestin (17-alpha hydroxyprogesterone caproate, ‘Makena’) for the prevention of PTB. However, FDA-approval for atosiban has been denied due to controversy concerning its effectiveness in clinical trials to delay PTB and improve neonatal outcomes. Also, FDA approval for Makena has been withdrawn due to the post marketing inability to verify any clinical benefit.

[0005] Preterm birth is also associated with high perinatal morbidity and mortality. Suppression of premature labor has not conventionally been associated with improved infantVU Ref.24027 / MBF Ref.093386-0031-WO01 outcomes. The conventional art does not show any direct health benefits to the neonate and the current therapies may have adverse side effects such as those listed above.

[0006] Thus, there remains a considerable unmet medical need for safe and effective treatments for or prevention of the onset of preterm labor and resulting PTB. Further, there remains a need for a novel method of treatment for or prevention of PTB that targets systems other than or in addition to smooth muscle. Moreover, beyond a therapy for improving the health of the mother giving birth, there needs to be an intervention for improving neonatal outcome for births occurring preterm. SUMMARY

[0007] In an aspect, the disclosure relates to a method for treating or preventing preterm labor or decreasing preterm birth in a pregnant subject in need thereof comprising administering to the subject a composition comprising a therapeutically effective amount of aprepitant or prodrug thereof. In an embodiment, the composition comprises a pharmaceutically acceptable carrier. In another embodiment, the prodrug is fosaprepitant.

[0008] In a further aspect, the disclosure relates to a method for treating or preventing preterm labor or decreasing preterm birth in a pregnant subject in need thereof comprising administering to the subject a composition comprising a therapeutically effective amount of ciclesonide. In an embodiment, the composition comprises a pharmaceutically acceptable carrier. In another embodiment, the composition further comprises administering atosiban to the subject. In another embodiment, the ciclesonide and the atosiban are administered to the subject at a ratio of about 11.8:1. In another embodiment, the ciclesonide and the atosiban are administered concomitantly. In another embodiment, the composition further comprises administering ivermectin to the subject. In another embodiment, the ciclesonide and the ivermectin are administered to the subject at a ratio of about 1.8:1. In another embodiment, the ciclesonide and the ivermectin are administered concomitantly.

[0009] Another aspect of the disclosure provides a method for treating or preventing preterm labor or decreasing preterm birth in a pregnant subject in need thereof comprising administering to the subject a composition comprising a therapeutically effective amount of ivermectin. In an embodiment, the ivermectin is administered in the form of a composition, wherein the composition comprises a pharmaceutically acceptable carrier. In another embodiment, the composition further comprises administering indomethacin to the subject. InVU Ref.24027 / MBF Ref.093386-0031-WO01 another embodiment, the ivermectin and the indomethacin are administered to the subject at a ratio of about 1:7.7. In another embodiment, the ivermectin and the indomethacin are administered concomitantly.

[0010] Another aspect of the disclosure provides a method for treating or preventing preterm labor or decreasing preterm birth in a pregnant subject in need thereof comprising administering to the subject a composition comprising a therapeutically effective amount of nelfinavir mesylate. In another embodiment, the composition comprises a pharmaceutically acceptable carrier. In another embodiment, the composition further comprises administering indomethacin to the subject. In another embodiment, the nelfinavir mesylate and the indomethacin are administered to the subject at a ratio of about 1:3. In another embodiment, the nelfinavir mesylate and the indomethacin are administered concomitantly. In another embodiment, the composition is administered orally, vaginally, rectally, subcutaneously, intraveneously, or by intramuscular injection. In another embodiment, the composition is administered once or more than once. In another embodiment, the composition is administered to the subject once or more than once per day. In another embodiment, the method prevents the onset of preterm labor. In another embodiment, the composition is administered after the onset of preterm labor. In another embodiment, the composition is administered before the onset of preterm labor. In another embodiment, the subject exhibits one or more of the following before administration of the composition: recurrent uterine contractions^ cervical dilation^ cervical effacement^ and a cervical change. In another embodiment, the composition is administered with a secondary treatment for preterm labor. In another embodiment, the secondary treatment for preterm labor comprises nifedipine, indomethacin, magnesium sulfate, terbutaline, Ritodrine, Atosiban, Barusiban, nitroglycerin, Makena, hydroxyprogesterone caproate, progesterone supplementation, or cervical cerclage. In another embodiment, the composition is administered repeatedly beginning about the 18th to 22nd week of gestation until about the 37th week of gestation. In another embodiment, the composition is administered repeatedly for about 14 to 19 weeks. In another embodiment, the method further comprises monitoring or confirming a reduction in effacement, uterine contractions, cervical dilation, or a combination thereof after the administration of the composition. In another embodiment, the composition prolongs gestation.

[0011] Another aspect of the disclosure provides a method for treating or preventing preterm labor or decreasing preterm birth in a pregnant subject in need thereof comprising administering to the subject a composition comprising a therapeutically effective amount ofVU Ref.24027 / MBF Ref.093386-0031-WO01 aprepritant, fosaprepitant, ciclesonide, atosiban, ivermectin, indomethacin, nelfinavir mesylate, or a combination thereof.

[0012] The disclosure provides for other aspects and embodiments that will be apparent in light of the following detailed description and accompanying figures. BRIEF DESCRIPTION OF THE DRAWINGS

[0013] FIGS.1A-M show identification of uterine-selective FDA-approved drugs with favorable in vitro therapeutic index and tocolytic potential. FIG.1A is a graph showing a systematic screening strategy for repurposing FDA-approved drugs for tocolytic use. FIG.1B is a graph showing real-time monitoring of Ca2+-mobilization from UT-myo cells, in which compounds are added (left panel), followed by OT (right panel). Hit-agonists are identified by compounds that induce Ca2+-mobilization, while hit-antagonists are identified as inhibitors of OT-induced Ca2+-mobilization from UT-myo cells. FIG.1C is a graph showing percent inhibition of OT-induced Ca2+-mobilization from the cells by the compounds (highlighted by grey), which were determined based on 3 mean absolute deviations above the average % inhibition from vehicle (DMSO) treated wells (hit selection threshold represented by dashed line). FIG.1D is a graph showing the average maximum % inhibition of each hit-antagonist (black and grey bars) after concentration-response analysis, grouped by their drug classes. Hit-antagonists were counter screened against VSMCs, yielding 12 uterine-selective drugs (highlighted grey bars) with ^ 5-fold greater Emaxor IC50values in myometrial cells. FIG.1E is a graph showing real- time monitoring of concentration-dependent inhibition of oxytocin-induced Ca2+-mobilization from UT-myo cells (left panel) and U46619-induced Ca2+-mobilization from aorta VSMCs (right panel) for Fosaprepitant Dimeglumine. FIG.1F is a graph showing percent inhibition of Ca2+- mobilization from the cells with increasing concentrations of Fosaprepitant Dimeglumine. FIG. 1G is a graph showing real-time monitoring of concentration-dependent inhibition of oxytocin- induced Ca2+-mobilization from UT-myo cells (left panel) and U46619-induced Ca2+-mobilization from aorta VSMCs (right panel) for Ciclesonide. FIG.1H is a graph showing percent inhibition of Ca2+-mobilization from the cells with increasing concentrations of Ciclesonide. FIG.1I is a graph showing real-time monitoring of concentration-dependent inhibition of oxytocin-induced Ca2+-mobilization from UT-myo cells (left panel) and U46619-induced Ca2+-mobilization from aorta VSMCs (right panel) for Nelfinavir Mesylate. FIG.1J is a graph showing percent inhibition of Ca2+-mobilization from the cells with increasing concentrations of Nelfinavir Mesylate. FIG. 1K is a graph showing real-time monitoring of concentration-dependent inhibition of oxytocin-VU Ref.24027 / MBF Ref.093386-0031-WO01 induced Ca2+-mobilization from UT-myo cells (left panel) and U46619-induced Ca2+-mobilization from aorta VSMCs (right panel) for Ivermectin. FIG.1L is a graph showing percent inhibition of Ca2+-mobilization from the cells with increasing concentrations of Ivermectin. FIG.1M is graphs showing representative CRCS from in vitro cell viability assays using myometrial (hTERT-HM), liver (HepG2), and kidney (RPTEC) cells after 72 hr incubation with fosaprepitant dimeglumine, ciclesonide, DMSO (vehicle control), or napabucasin (positive control, known-toxic compound). Non-linear regression was used to fit the data (mean ± SD) and calculate IC50 values. These two drugs had favorable in vitro therapeutic indexes ^5, which were calculated as the ratio between IC50 values obtained from the Ca2+-mobilization and cell viability assays. ***Pௗ<ௗ0.001,

[0014] FIGS.2A-K show identification of synergistic combinations of uterine-selective FDA- approved drugs with favorable in vitro therapeutic index and tocolytic potential. FIG.2A is a flowchart showing an exemplary systematic screening strategy to identify synergistic combinations of FDA-approved drugs. FIG.2B is a matrix showing two-fold titrations above and below the IC50 value of each uterine-selective hit-antagonist and 3 current tocolytic drugs (atosiban, indomethacin, and nifedipine) that were tested in combination using a high- throughput Ca2+-mobilization assay and primary myometrial cells. FIG.2C is a Venn diagram showing percent response data for each compound concentration that was analyzed using online (Combenefit) and in-house built (MuSyc) tools to provide synergy scores. The Venn diagram shows the number of overlapping and non-overlapping synergistic drug combinations identified by each tool. FIG.2D shows a representative bliss-independence heat map of drug combination ratios of atosiban and ciclesonide. FIG.2E shows a representative bliss- independence heat map of drug combination ratios of ivermectin and ciclesonide. FIG.2F shows a representative bliss-independence heat map of drug combination ratios of indomethacin and nelfinavir mesylate. FIG.2G shows a representative bliss-independence heat map of drug combination ratios of indomethacin and ivermectin. FIGS.2H-K are graphs showing percent inhibition of Ca2+-mobilization from the cells with increasing concentrations of the drug combinations tested at their fixed-ratios identified as synergistic using the combination high-throughput screen. FIG.2H is a graph showing representative CRCs of synergistic fixed- ratios of atosiban and ciclesonide and their respective concentration for each. FIG.2I is a graph showing representative CRCs of synergistic fixed-ratios of ivermectin and ciclesonide and their respective concentration for each. FIG.2J is a graph showing representative CRCs of synergistic fixed-ratios of indomethacin and ivermectin and their respective concentration forVU Ref.24027 / MBF Ref.093386-0031-WO01 each. FIG.2K is a graph showing representative CRCs of synergistic fixed-ratios of indomethacin and nelfinavir mesylate and their respective concentration for each. For FIGS. 2H-K, non-linear regression was used to fit the data (mean ± SD) and calculate the IC50 and Emax values. A 2-way ANOVA with a post-hoc Fisher’s LSD test was used to compare the % inhibition at each concentration.

[0015] FIGS.3A-L show effects of drug combinations on cell viability. FIG.3A is a graph showing the effect of fixed-ratio drug combination Ciclesonide + Atosiban (11.8:1) and the individual drugs at varying concentrations on cell viability of hTERT-HM cells. FIG.3B is a graph showing the effect of drug combination Ciclesonide + Atosiban (11.8:1) and the individual drugs at varying concentrations on cell viability of HepG2 cells. FIG.3C is a graph showing the effect of drug combination Ciclesonide + Atosiban (11.8:1) and the individual drugs at varying concentrations on cell viability of RPTEC cells. FIG.3D is a graph showing the effect of fixed- ratio drug combination Ciclesonide + Ivermectin (1.8:1) and the individual drugs at varying concentrations on cell viability of hTERT-HM cells. FIG.3E is a graph showing the effect of drug combination Ciclesonide + Ivermectin (1.8:1) and the individual drugs at varying concentrations on cell viability of HepG2 cells. FIG.3F is a graph showing the effect of drug combination Ciclesonide + Ivermectin (1.8:1) and the individual drugs at varying concentrations on cell viability of RPTEC cells. FIG.3G is a graph showing the effect of fixed-ratio drug combination Ivermectin + Indomethacin (1:7.7) and the individual drugs at varying concentrations on cell viability of hTERT-HM cells. FIG.3H is a graph showing the effect of drug combination Ivermectin + Indomethacin (1:7.7) and the individual drugs at varying concentrations on cell viability of HepG2 cells. FIG.3I is a graph showing the effect of drug combination Ivermectin + Indomethacin (1:7.7) and the individual drugs at varying concentrations on cell viability of RPTEC cells. FIG.3J is a graph showing the effect of fixed- ratio drug combination Nelfinavir Mesylate + Indomethacin (1:3) and the individual drugs at varying concentrations on cell viability of hTERT-HM cells. FIG.3K is a graph showing the effect of drug combination Nelfinavir Mesylate + Indomethacin (1:3) and the individual drugs at varying concentrations on cell viability of HepG2 cells. FIG.3L is a graph showing the effect of drug combination Nelfinavir Mesylate + Indomethacin (1:3) and the individual drugs at varying concentrations on cell viability of RPTEC cells.

[0016] FIGS.4A-J show ex vivo tocolytic efficacy and potency of single hit-antagonists identified as uterine-selective with favorable in vitro therapeutic index. FIG.4A is representative recordings of isometric spontaneous contractility of human myometrial tissue before the additionVU Ref.24027 / MBF Ref.093386-0031-WO01 of increasing concentrations (1 nM - 100 ^M) of vehicle, ciclesonide, fosaprepitant, aprepitant, anidulafungin, fenticonazole, ivermectin, nebivolol, nelfinavir mesylate, and tiratricol. FIG.4B is representative tracings of fetal mouse ductus arteriosus (DA) tone (measured by lumen diameter) at baseline (“B”) and after the addition of increasing concentrations (1ௗnm - 100ௗ^M) of vehicle control, ciclesonide, fosaprepitant dimeglumine, and aprepitant. For FIGS.4C-F, contractile recordings from FIG.4A were analyzed for area under the contractile curve and non- linear regression was used to fit the % response data (mean ± SD) to determine the effect of compounds on myometrial contractility and to calculate EC50 and Emax. A 2-way ANOVA with a post-hoc Tukey analysis was used to compare the % inhibition at each concentration relative to vehicle control. FIG.4C is a graph showing percent response for contractile recordings with a vehicle control and ciclesonide. FIG.4D is a graph showing percent response for contractile recordings with a vehicle control, fosaprepitant, and aprepitant. FIG.4E is a graph showing percent response for contractile recordings with a vehicle control, anidulafungin, fenticonazole, and ivermectin. FIG.4F is a graph showing percent response for contractile recordings with a vehicle control, nebivolol, nelfinavir mesylate, and tiratricol. For FIGS.4G-J, vasoconstrictive effects of drugs were tested using cannulated term gestation mouse fetal ductus arteriosus in an ex vivo pressure myography assay. Ductus tone at baseline and after increasing concentrations (1 nM - 100 ^M) of either vehicle or drugs was analyzed for % change from baseline (“BL”) lumen diameter, followed by non-linear regression to fit the data. FIG.4G is a graph showing percent change from BL with a vehicle control and ciclesonide. FIG.4H is a graph showing percent change from BL with a vehicle control, fosaprepitant, and aprepitant. FIG.4I is a graph showing percent change from BL with a vehicle control, anidulafungin, fenticonazole, and ivermectin. FIG.4J is a graph showing percent change from BL with a vehicle control, nebivolol, nelfinavir mesylate, and tiratricol.

[0017] FIGS.5A-E show ex vivo tocolytic efficacy and potency of synergistic hit-antagonists identified as uterine-selective with favorable in vitro therapeutic index. Area under the contractile curve was analyzed. Non-linear regression was used to fit the % response data (mean ± SD) and to calculate EC50 and Emax. A 2-way ANOVA with a post-hoc Tukey analysis was used to compare the % inhibition at each concentration. Asterisks indicate significantly different percent response between drug combinations and their single agents. FIG.5A shows representative recordings (top) of isometric spontaneous contractility of human myometrial tissue before the addition of increasing concentrations (1 nM - 100 ^M) and concentration–response curve (CRC) analysis (bottom) for ciclesonide alone, atosiban alone,VU Ref.24027 / MBF Ref.093386-0031-WO01 and the fixed-ratio drug combination of ciclesonide and atosiban. FIG.5B shows representative recordings (top) of isometric spontaneous contractility of human myometrial tissue before the addition of increasing concentrations (1 nM - 100 ^M) and CRC analysis (bottom) for ciclesonide alone, ivermectin alone, and the fixed-ratio drug combination of ciclesonide and ivermectin. FIG.5C shows representative recordings (top) of isometric spontaneous contractility of human myometrial tissue before the addition of increasing concentrations (1 nM - 100 ^M) and CRC analysis (bottom) for ivermectin alone, indomethacin alone, and the fixed- ratio drug combination of ivermectin and indomethacin. FIG.5D shows representative recordings (top) of isometric spontaneous contractility of human myometrial tissue before the addition of increasing concentrations (1 nM - 100 ^M) and CRC analysis (bottom) for nelfinavir mesylate alone, indomethacin alone, and the fixed-ratio drug combination of nelfinavir mesylate and indomethacin. FIG.5E is representative tracings of fetal mouse ductus arteriosus (DA) tone (measured by lumen diameter) at baseline (“B”) and after the addition of increasing concentrations (1ௗnm - 100ௗ^M) of indomethacin alone, indomethacin + ivermectin, and indomethacin + nelfinavir mesylate (“NM”^ top). Vasoconstrictive effects of drugs were tested using cannulated term gestation mouse fetal ductus arteriosus in an ex vivo pressure myography assay. Ductus tone at baseline and after increasing concentrations (1 nM - 100 ^M) of either vehicle or drugs was analyzed for % change from baseline (“BL”) lumen diameter, followed by non-linear regression to fit the data. FIG.5E is also a graph showing percent change from BL with a of indomethacin alone, indomethacin + ivermectin, and indomethacin + nelfinavir mesylate (bottom).

[0018] FIGS.6A-D show in vivo efficacy of single FDA-approved drugs to delay delivery and improve neonatal outcomes in a preclinical mouse model of induced PTL. FIG.6A is a schematic showing an exemplary experimental timeline for induction of preterm labor and birth using mifepristone (MIF) on day 15 of pregnancy, followed by s.c. administration of either vehicle or single-drugs in mice. Dosing strategies were optimized to yield maximum maternal and fetal outcomes. Dose-response analysis of aprepitant (30, 50, 70 mg / kg) starting 5 hrs post-MIF followed by injections every 12 hours until gd18 am. FIG.6B is a graph showing timing of delivery for untreated (control), vehicle (control)-treated, and aprepitant-treated MIF- induced PTL mice. Dotted line represents PTB, defined as 24 hrs prior to the average timing of term delivery of untreated mice. Term birth rates and pup viability (%) are provided for each group. A one-way ANOVA with an uncorrected Fisher’s LSD test was used to compare aprepitant-treatment to untreated and vehicle-treated controls. FIG.6C is images of newbornVU Ref.24027 / MBF Ref.093386-0031-WO01 pups that were imaged before and after assessment of skin maturation using Toluidine blue dye. FIG.6D is images of H&E-stained sections of newborn pup lungs and a graph showing morphometric analysis of airspace volume density (% ASVD) of the images. A one-way ANOVA with a posthoc Sidak test was used to compare treatment to untreated and vehicle- treated controls.Preterm gd16 pup and lung images are from vehicle-treated MIF mice. Term P0 pup and lung images are from untreated (control) mice.

[0019] FIGS.7A-D show in vivo efficacy of single FDA-approved drugs to delay delivery and improve neonatal outcomes in a preclinical mouse model of induced PTL. FIG.7A is a schematic showing an exemplary experimental timeline for induction of preterm labor and birth using mifepristone (MIF) on day 15 of pregnancy, followed by s.c. administration of either vehicle or single-drugs in mice. Dosing strategies were optimized to yield maximum maternal and fetal outcomes. Ciclesonide was tested using 5 dosing strategies (DS). First, 30 mg / kg was given 5hrs post-MIF followed by once-daily doses from gd16-18. Second, the last dose of ciclesonide on gd18 was eliminated. Third, various doses (7.5, 15, 30, 40 or 50 mg / kg) were tested at 5 hrs and 17 hrs (i.e. gd16 am) post-MIF. In the 4th and 5th DS, a loading dose (20 or 30 mg / kg) was given 5 hrs post-MIF, followed by once daily am injections of a loading dose (1.8 mg / kg). FIG.7B is a graph showing timing of delivery for untreated (control), vehicle (control)- treated, and ciclesonide-treated (dosing strategies 1-5) MIF-induced PTL mice. A one-way ANOVA with an uncorrected Fisher’s LSD test was used to compare ciclesonide-treatment to untreated and vehicle-treated controls. FIG.7C is images of newborn pups that were imaged before and after assessment of skin maturation using Toluidine blue dye. FIG.7D is images of H&E-stained sections of newborn pup lungs and a graph showing morphometric analysis of airspace volume density (% ASVD) of the images. A one-way ANOVA with a posthoc Sidak test was used to compare treatment to untreated and vehicle-treated controls. ****Pௗ<ௗ0.0001. Preterm gd16 pup and lung images are from vehicle-treated MIF mice. Term P0 pup and lung images are from untreated (control) mice.

[0020] FIGS.8A-D show in vivo efficacy of ciclesonide drug combinations to delay delivery and improve neonatal outcomes in a preclinical mouse model of induced PTL. FIG.8A is a schematic showing an exemplary experimental timeline for induction of preterm labor and birth using mifepristone (MIF) on day 15 of pregnancy, followed by s.c. administration of either vehicle, syngergistic fixed-ratio of drug combinations containing ciclesonide + atosiban or ciclesonide + ivermectin. Single-agent controls were tested at their respective doses in the combination. FIG.8B is a graph showing timing of delivery for untreated (control), vehicleVU Ref.24027 / MBF Ref.093386-0031-WO01 (control)- and ciclesonide combinations and their respective single-agent control mice. Dotted line represents PTB, defined as 24 hrs before the average timing of term delivery of untreated mice. Term birth rates and pup viability (%) are provided for each group. A one-way ANOVA with a posthoc Sidak test was used to compare single and combination treatments to untreated controls, as well as combination vs single drug treatment. FIG.8C is images of newborn pups that were imaged before and after assessment of skin maturation using Toluidine blue dye. FIG.8D is images of H&E-stained sections of newborn pup lungs and a graph showing morphometric analysis of airspace volume density (% ASVD) from the images. A one-way ANOVA with a posthoc Sidak test was used to compare treatment to untreated and vehicle- treated controls. ****Pௗ<ௗ0.0001. Preterm gd16 pup and lung images are from vehicle-treated MIF mice. Term P0 pup and lung images are from untreated (control) mice.

[0021] FIGS.9A-E show in vivo efficacy of indomethacin drug combinations to delay delivery and improve neonatal outcomes in a preclinical mouse model of induced PTL. FIG.9A is a schematic showing an exemplary experimental timeline for induction of preterm labor and birth using mifepristone (MIF) on day 15 of pregnancy, followed by s.c. administration of either vehicle, synergistic fixed-ratio of drug combinations containing indomethacin + ivermectin or indomethacin + nelfinavir mesylate. Single-agent controls were tested at their respective doses in the combination. FIG.9B is a graph showing timing of delivery for untreated (control), vehicle (control)- and indomethacin combinations and their respective single-agent control mice. Dotted line represents PTB, defined as 24 hrs before the average timing of term delivery of untreated mice. Term birth rates and pup viability (%) are provided for each group. A one-way ANOVA with a posthoc Sidak test was used to compare single and combination treatments to untreated controls, as well as combination vs single drug treatment. FIG.9C is images showing newborn pups that were imaged before and after assessment of skin maturation using Toluidine blue dye. FIG.9D is images showing H&E-stained sections of newborn pup lungs and a graph showing morphometric analysis of airspace volume density (% ASVD) of the images. A one- way ANOVA with a posthoc Sidak test was used to compare single and combination treatments to untreated and vehicle-treated controls, as well as combination vs single drug treatment. ****Pௗ<ௗ0.0001. Preterm gd16 pup and lung images are from vehicle-treated MIF mice. Term P0 pup and lung images are from untreated (control) mice. FIG.9E is representative images showing ductus arteriosus (DA, indicated by arrow) closure at 4ௗh of age in a term P0 pup born to an untreated (control) dam and persistent patency of the DA in a P0 pup born to an indomethacin (3.7 mg / kg)-treated dam^ and a graph showing percent DA patency. AO = aorta,VU Ref.24027 / MBF Ref.093386-0031-WO01 PA = pulmonary artery. A one-way ANOVA with a posthoc Sidak test was used to compare treatment groups. DETAILED DESCRIPTION

[0022] A total of 52 drugs were found to inhibit myometrial calcium (Ca2+)-mobilization from varying drug classes, including: antifungal agents, anti-infective agents, and anti-asthmatic agents / leukotriene antagonists. To limit off-target effects of current tocolytics, hit-antagonists against various vascular smooth muscle cells were counter-screened, resulting in 10 uterine- selective drugs that lack reported reproductive toxicity. Of these, 2 drugs (Fosaprepitant Dimeglumine and Ciclesonide) displayed a favorable in vitro therapeutic index (TI>5) and tocolytic efficacy in an ex vivo contractility assay using human myometrial tissue but lacked ex vivo effects on term fetal murine ductus arteriosus tone. Next, the synergistic ability of hit- antagonists was analyzed by performing a combination high-throughput screen using a Ca2+- mobilization assay and primary myometrial cells. Four drug combinations (ciclesonide+atosiban, ciclesonide+ivermectin, indomethacin+nelfinavir mesylate and indomethacin+ivermectin) displayed synergy with an in vitro TI>5, and improved ex vivo tocolytic potency and / or efficacy. Finally, it was demonstrated herein that the in vivo efficacy of the above two single agents (aprepitant, the parent drug of fosaprepitant, and ciclesonide) and four drug combinations to delay delivery until term in a mouse model of mifepristone-induced preterm labor, as well as improve neonatal viability and developmental maturation of skin and lungs. 1. Definitions

[0023] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art. The meaning and scope of the terms should be clear. In case of conflict, the present document, including definitions, take precedent over any dictionary or extrinsic definition. Further, unless otherwise required by context, singular terms shall include pluralities and plural terms shall include the singular. Preferred methods and materials are described below, although methods and materials similar or equivalent to those described herein can be used in practice or testing of the present invention. All publications, patent applications, patents and other references mentioned herein are incorporated by reference in their entirety. The materials, methods, and examples disclosed herein are illustrative only and not intended to be limiting.VU Ref.24027 / MBF Ref.093386-0031-WO01

[0024] The terms “comprise(s),” “include(s),” “having,” “has,” “can,” “contain(s),” and variants thereof, as used herein, are intended to be open-ended transitional phrases, terms, or words that do not preclude the possibility of additional acts or structures. The singular forms “a,” “and,” and “the” include plural references unless the context clearly dictates otherwise. The present disclosure also contemplates other embodiments “comprising,” “consisting of,” and “consisting essentially of,” the embodiments or elements presented herein, whether explicitly set forth or not.

[0025] For the recitation of numeric ranges herein, each intervening number there between with the same degree of precision is explicitly contemplated. For example, for the range of 6-9, the numbers 7 and 8 are contemplated in addition to 6 and 9, and for the range 6.0-7.0, the number 6.0, 6.1, 6.2, 6.3, 6.4, 6.5, 6.6, 6.7, 6.8, 6.9, and 7.0 are explicitly contemplated.

[0026] The term “about” or “approximately” as used herein as applied to one or more values of interest, refers to a value that is similar to a stated reference value, or within an acceptable error range for the particular value as determined by one of ordinary skill in the art, which will depend in part on how the value is measured or determined, such as the limitations of the measurement system. In certain aspects, the term “about” refers to a range of values that fall within 20%, 19%, 18%, 17%, 16%, 15%, 14%, 13%, 12%, 11%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, or less in either direction (greater than or less than) of the stated reference value unless otherwise stated or otherwise evident from the context (except where such number would exceed 100% of a possible value). Alternatively, “about” can mean within 3 or more than 3 standard deviations, per the practice in the art. Alternatively, such as with respect to biological systems or processes, the term “about” can mean within an order of magnitude, in some embodiments within 5-fold, and in some embodiments within 2-fold, of a value.

[0027] The terms “active ingredient” or “active pharmaceutical ingredient” as used herein refer to a pharmaceutical agent, active ingredient, compound, or substance, compositions, or mixtures thereof, that provide a pharmacological, often beneficial, effect.

[0028] The terms “control,” “reference level,” and “reference” are used herein interchangeably. The reference level may be a predetermined value or range, which is employed as a benchmark against which to assess the measured result. “Control group” as used herein refers to a group of control subjects. The predetermined level may be a cutoff value from a control group. The predetermined level may be an average from a control group.VU Ref.24027 / MBF Ref.093386-0031-WO01 Cutoff values (or predetermined cutoff values) may be determined by Adaptive Index Model (AIM) methodology. Cutoff values (or predetermined cutoff values) may be determined by a receiver operating curve (ROC) analysis from biological samples of the patient group. ROC analysis, as generally known in the biological arts, is a determination of the ability of a test to discriminate one condition from another, e.g., to determine the performance of each marker in identifying a patient having CRC. A description of ROC analysis is provided in P.J. Heagerty et al. (Biometrics 2000, 56, 337-44), the disclosure of which is hereby incorporated by reference in its entirety. Alternatively, cutoff values may be determined by a quartile analysis of biological samples of a patient group. For example, a cutoff value may be determined by selecting a value that corresponds to any value in the 25th-75th percentile range, in some embodiments a value that corresponds to the 25th percentile, the 50th percentile or the 75th percentile, and in some embodiments the 75th percentile. Such statistical analyses may be performed using any method known in the art and can be implemented through any number of commercially available software packages (e.g., from Analyse-it Software Ltd., Leeds, UK^ StataCorp LP, College Station, TX^ SAS Institute Inc., Cary, NC.). The healthy or normal levels or ranges may be defined in accordance with standard practice. A control may be a subject or cell without a drug as detailed herein. A control may be a subject, or a sample therefrom, whose disease state is known. The subject, or sample therefrom, may be healthy, diseased, diseased prior to treatment, diseased during treatment, or diseased after treatment, or a combination thereof.

[0029] The term “dose” as used herein denotes any form of the active ingredient formulation or composition that contains an amount sufficient to produce a therapeutic effect with at least a single administration. “Formulation” and “compound” are used interchangeably herein.

[0030] The term “dosage” as used herein refers to the administering of a specific amount, number, and frequency of doses over a specified period of time, typically 1 day.

[0031] As used herein, “neonatal” encompasses children about 6 months of age or less, preferably about 3 months of age or less, more preferably about 2 months of age or less, and even more preferably about 1 month of age or less. In certain embodiments, neonatal is used to encompass perinatal, preferably the period after birth.

[0032] As used herein, “neonatal outcome,” interchangeably referred to herein as “neonatal health,” is measured by mortality and morbidity incidences, prevalence, and clinical states. For example, health may be directly correlated to infant weight. Health may also be inverselyVU Ref.24027 / MBF Ref.093386-0031-WO01 related to incidence of special care admission (i.e., neonatal intensive care unit (NICU)) at birth, length of neonatal hospital stay at birth, incidence of respiratory distress syndrome (RDS), incidence and severity of intraventricular hemorrhage, incidence and severity of patent ductus arteriosus, incidence of necrotizing enterocolitis, incidence of sepsis, and incidence of neonatal death. Other measures of morbidity may be used by those skilled in the art.

[0033] As used herein, the terms “preterm” or “premature” generally describes human gestation resulting in birth prior to 37 weeks, also known as “preterm birth” or “PTB”. Accordingly, “preterm” or “preterm birth” covers births occurring less than 35 weeks or less than or equal to 32 weeks of gestation.

[0034] As used herein, the terms “preterm labor” or “premature labor” or “PTL” includes dilation and / or effacement of the cervix, which is detected by digital examination, associated with persistent uterine contractions before 37 weeks of gestation. For example, preterm labor may be defined as 6 or more uterine contractions per hour accompanied by documented cervical change, cervical dilation greater than 2 cm, cervical effacement greater than 80%, or documented change in cervical effacement greater than 50%.

[0035] The term “prophylaxis” refers to preventing or reducing the progression of a disorder, either to a statistically significant degree or to a degree detectable to one skilled in the art.

[0036] “Sample” or “test sample” as used herein can mean any sample in which the presence and / or level of a target is to be detected or determined or any sample comprising a drug as detailed herein. Samples may include liquids, solutions, emulsions, or suspensions. Samples may include a medical sample. Samples may include any biological fluid or tissue, such as blood, whole blood, fractions of blood such as plasma and serum, muscle, interstitial fluid, sweat, saliva, urine, tears, synovial fluid, bone marrow, cerebrospinal fluid, nasal secretions, sputum, amniotic fluid, bronchoalveolar lavage fluid, gastric lavage, emesis, fecal matter, lung tissue, peripheral blood mononuclear cells, total white blood cells, lymph node cells, spleen cells, tonsil cells, cancer cells, tumor cells, bile, digestive fluid, skin, or combinations thereof. In some embodiments, the sample comprises an aliquot. In other embodiments, the sample comprises a biological fluid. Samples can be obtained by any means known in the art. The sample can be used directly as obtained from a patient or can be pre- treated, such as by filtration, distillation, extraction, concentration, centrifugation, inactivation ofVU Ref.24027 / MBF Ref.093386-0031-WO01 interfering components, addition of reagents, and the like, to modify the character of the sample in some manner as discussed herein or otherwise as is known in the art.

[0037] “Subject,” “study participant,” “participant,” and “patient” as used herein interchangeably refers to any vertebrate, including, but not limited to, a mammal that wants or is in need of the herein described compounds, compositions, or methods. The subject may be a human or a non-human. The subject may be a vertebrate. The subject may be a mammal. The mammal may be a primate or a non-primate. The mammal can be a non-primate such as, for example, cow, pig, camel, llama, hedgehog, anteater, platypus, elephant, alpaca, horse, goat, rabbit, sheep, hamsters, guinea pig, cat, dog, rat, and mouse. The mammal can be a primate such as a human. The mammal can be a non-human primate such as, for example, monkey, cynomolgus monkey, rhesus monkey, chimpanzee, gorilla, orangutan, and gibbon. The subject may be of any age or stage of development, such as, for example, an adult, an adolescent, or an infant. The subject may be male. The subject may be female. In some embodiments, the subject is pregnant. The subject may be undergoing other forms of treatment.

[0038] “Treatment” or “treating” when referring to protection of a subject from a disease or disorder, means suppressing, repressing, reversing, alleviating, ameliorating, or inhibiting the progress of disease or disorder, or completely eliminating a disease or disorder. A treatment may be either performed in an acute or chronic way. The term also refers to reducing the severity of a disease or a disorder or symptoms associated with such disease or disorder prior to affliction with the disease or disorder. Preventing the disease or disorder involves administering a composition or compound of the present disclosure to a subject prior to onset of the disease or disorder. Suppressing the disease or disorder involves administering a composition or compound of the present disclosure to a subject after induction of the disease or disorder but before its clinical appearance. Repressing or ameliorating the disease or disorder involves administering a composition or compound of the present disclosure to a subject after clinical appearance of the disease or disorder. 2. Compounds for Treating or Preventing Preterm Labor or Decreasing Preterm Birth

[0039] Provided herein are methods for treating or preventing preterm labor or decreasing preterm birth in a pregnant subject in need thereof using aprepitant or a prodrug thereof. Aprepitant may also be named as 2-(R)-(1-(R)-(3,5-bis(trifluoromethyl)phenyl)ethoxy)-3-(S)-(4-VU Ref.24027 / MBF Ref.093386-0031-WO01 fluoro)phenyl-4-(3-(5-oxo-1H,4H-1,2,4-triazolo)methylmorpholine and is described in U.S. Patent No.5,719,147 and U.S. Patent No.8,258,132, which are fully incorporated herein by reference. Aprepitant is a Food and Drug Administration (FDA)-approved compound that is an orally available antiemetic agent that is traditionally used to prevent postoperative or cancer chemotherapy-related nausea and vomiting. Aprepitant has the following structural formula, Formula (I):

[0040] The prodrug of aprepitant may be fosaprepitant. Fosaprepitant may also be named as fosaprepitant dimeglumine or EMEND® or 1-Deoxy-1-(methylamino)-D-glucitol[3-[[(2R,3S)-2- [(1R)-1-[3,5-bis(trifluoromethyl)phenyl]ethoxy]-3-(4-fluorophenyl)-4-morpholinyl]methyl]-2,5- dihydro-5-oxo-1H-1,2,4-triazol-1-yl]phosphonate. Fosaprepitant is described in U.S. Patent No. 5,512,570 and U.S. Patent No.5,691,336, which are fully incorporated herein by reference. Fosaprepitant is an FDA-approved compound that is a phosphorylated analog of aprepitant that is traditionally used to prevent postoperative or cancer chemotherapy-related nausea and vomiting. Aprepitant has the following structural formula, Formula (II):VU Ref.24027 / MBF Ref.093386-0031-WO01

[0041] Further provided herein are methods for treating or preventing preterm labor or decreasing preterm birth in a pregnant subject in need thereof using ciclesonide. Ciclesonide may also be named as [11ȕ,16Į(R)]-16,17-[(Cyclohexylmethylen)bis(oxy)]-11-hydroxy-21-(2- methyl-1-oxopropoxy)pregna-1,4-dien-3,20-dion and is described in U.S. Patent No.5,482,934 and U.S. Patent No.8,371,292, which are fully incorporated herein by reference. Ciclesonide is an FDA-approved compound that is a glucocorticoid traditionally used in the symptomatic relief of nasal symptoms associated with seasonal and perennial allergic rhinitis. Ciclesonide has the following structural formula, Formula (III):

[0042] Further provided herein are methods for treating or preventing preterm labor or decreasing preterm birth in a pregnant subject in need thereof using ivermectin. Ivermectin is a mixture of two avermectins, comprising roughly 90% 5-O-demethyl-22,23-dihydroavermectin A1a(22,23-dihydroavermectin B1a) (“B1a”) and 10% 5-O-demethyl-25-de(1-methylpropyl)-22,23- dihydro--25-(1-methylethyl)avermectin A1a(22,23-dihydroavermectin B1b) (“B1b”) and is described in U.S. Patent No.4,199,569, which is fully incorporated herein by reference.VU Ref.24027 / MBF Ref.093386-0031-WO01 Ivermectin is an FDA-approved compound that is an anti-parasitic traditionally used to treat head lice, onchocerciasis, strongyloidiasis, ascariasis, trichuriasis, and enterobiasis. Ivermectin has the following structural formula, Formula (IV), wherein R is C2H5for B1aor CH3for B1b:

[0043] Further provided herein are methods for treating or preventing preterm labor or decreasing preterm birth in a pregnant subject in need thereof using nelfinavir mesylate. Nelfinavir mesylate may also be named as [3S-(3R,4aR,8aR,2ƍS,3ƍS)]-2-{2ƍ-hydroxy-3ƍ- [(phenylthio)methyl]-4ƍ-aza-5ƍ-oxo-5ƍ-(2Ǝ-methyl-3Ǝ-hydroxy phenyl)pentyl}decahydroiso- quinoline-3-N-tertbutylcarboxamide mesylate and is described in U.S. Patent No.5,484,926, which is fully incorporated herein by reference. Nelfinavir mesylate is an FDA-approved compound that is an HIV-protease inhibitor and is traditionally used as a treatment for HIV- infected individuals. Nelfinavir mesylate has the following structural formula, Formula (V):VU Ref.24027 / MBF Ref.093386-0031-WO01

[0044] Atosiban is an inhibitor of the hormones oxytocin and vasopressin and a known tocolytic that is used for suppressing labor in a pregnant subject. Atosiban may also be named as 1-(3-Mercaptopropanoic acid)-2-(O-ethyl-D-tyrosine)-4-L-threonine-8-L-ornithineoxytocin and is described in U.S. Patent No.4,504,469, which is fully incorporated herein by reference. Atosiban is not an FDA-approved compound due to the high number of infant deaths from its use. Atosiban has the following structural formula, Formula (VI):VU Ref.24027 / MBF Ref.093386-0031-WO01

[0045] Indomethacin is a nonsteroidal anti-inflammatory drug that is a prostaglandin inhibitor used off-label for tocolysis which is achieved by reversibly binding to cyclooxygenase. Indomethacin is an FDA-approved compound that is typically used to manage acute pain, rheumatoid arthritis, ankylosing spondylitis, osteoarthritis, bursitis, gouty arthritis, and patent ductus arteriosus. Indomethacin may also be named as 2-(1-(4-Chlorobenzoyl)-5-methoxy-2- methyl-1H-indol-3-yl)acetic acid and is described in U.S. Patent No.3,161,654, which is fully incorporated herein by reference. Indomethacin has the following structural formula, Formula (VII):VU Ref.24027 / MBF Ref.093386-0031-WO01

[0046] Provided herein are methods for treating or preventing preterm labor or decreasing preterm birth in a pregnant subject in need thereof using anidulafungin. Anidulafungin may also be named as N-{(2R,6S,9S,11R,12R,14aS,15S,16S,20S,23S,25aS)-23-[(1S,2S)-1,2-Dihydroxy- 2-(4-hydroxyphenyl)ethyl]-2,11,12,15-tetrahydroxy-6,20-bis[(1R)-1-hydroxyethyl]-16-methyl- 5,8,14,19,22,25-hexaoxotetracosahydr o-1H-dipyrrolo[2,1-c:2ƍ,1ƍ- l][1,4,7,10,13,16]hexaazacyclohenicosin-9-yl}-4Ǝ-(pentyloxy)-1,1ƍ:4ƍ,1Ǝ-terphenyl-4-carboxamide and is described in U.S. Patent No.5,965,525, U.S. Patent No.6,384,013, and U.S. Patent No. 6,743,777, which are fully incorporated herein by reference. Anidulafungin is a Food and Drug Administration (FDA)-approved compound that is an intravenously administered antifungal agent that is traditionally used to treat invasive candidiasis. Anidulafungin has the following structural formula, Formula (VIII):

[0047] Provided herein are methods for treating or preventing preterm labor or decreasing preterm birth in a pregnant subject in need thereof using conivaptan. Conivaptan may also be named as N-{4-[(2-Methyl-4,5-dihydroimidazo[4,5-d][1]benzazepin-6(1H)-yl)carbonyl]phenyl}-2- biphenylcarboxamide and is described in U.S. Patent No.5,723,606, which is fully incorporated herein by reference. Conivaptan is a Food and Drug Administration (FDA)-approved compound that is an intravenously administered vasopressin receptor antagonist that is traditionally used to treat hyponatremia (low blood sodium levels) caused by syndrome of inappropriate antidiuretic hormone (SIADH). Conivaptan has the following structural formula, Formula (IX):VU Ref.24027 / MBF Ref.093386-0031-WO01

[0048] Provided herein are methods for treating or preventing preterm labor or decreasing preterm birth in a pregnant subject in need thereof using benzbromarone. Benzbromarone may also be named as (3,5-Dibromo-4-hydroxyphenyl)(2-ethyl-1-benzofuran-3-yl)methanone and is described in U.S. Patent No.8,367,843, which is fully incorporated herein by reference. Benzbromarone is a nonpurine xanthine oxidase inhibitor that is traditionally used for the treatment of gout, but it is not FDA approved because of concerns over reports of acute liver injury and death. Benzbromarone has the following structural formula, Formula (X):

[0049] Provided herein are methods for treating or preventing preterm labor or decreasing preterm birth in a pregnant subject in need thereof using tiratricol. Tiratricol may also be named as [4-(4-Hydroxy-3-iodophenoxy)-3,5-diiodophenyl]acetic acid and is described in U.S. Patent No.2,970,165, which is fully incorporated herein by reference. Tiratricol is a thyroid hormone analogue that has been indicated in the management of thyroid hormone resistance syndrome, hyperlipidemia, localized lipodystrophy, obesity, and treatment of Allan-Herndon-Dudley Syndrome. Tiratricol has the following structural formula, Formula (XI):VU Ref.24027 / MBF Ref.093386-0031-WO013. Pharmaceutical Compositions

[0050] Further provided herein are pharmaceutical compositions comprising the above- described compounds or pharmaceutically acceptable salts thereof or prodrugs thereof. The pharmaceutical compositions may comprise therapeutically effective amounts of aprepitant, fosaprepitant, ciclesonide, ivermectin, nelfinavir mesylate, atosiban, indomethacin, anidulafungin, conivaptan, benzbromarone, tiratricol, or various combinations thereof. In some embodiments, the pharmaceutical compositions may comprise therapeutically effective amounts of aprepitant, fosaprepitant, ciclesonide, ivermectin, nelfinavir mesylate, atosiban, indomethacin, or various combinations thereof. The pharmaceutical composition may further comprise a pharmaceutically acceptable carrier.

[0051] Although anidulafungin, conivaptan, benzbromarone, and tiratricol were not uterine- selective or had an unfavorable therapeutic index, a pharmaceutical composition described herein may comprise one or more of anidulafungin, conivaptan, benzbromarone, and tiratricol. In some embodiments, to improve uterine-selective targeting and increase the therapeutic index of anidulafungin, conivaptan, benzbromarone, tiratricol, or various combinations thereof, the pharmaceutical composition comprising one or more of anidulafungin, conivaptan, benzbromarone, and tiratricol may be comprised by a pharmaceutically acceptable carrier such as a nanoparticle, drug conjugate, and the like.

[0052] Compounds or pharmaceutically acceptable salts thereof as detailed herein, or at least one component thereof, may be formulated into pharmaceutical compositions in accordance with standard techniques well known to those skilled in the pharmaceutical art. The pharmaceutical compositions can be formulated according to the mode of administration to be used. In cases where pharmaceutical compositions are injectable pharmaceutical compositions, they are sterile, pyrogen free, and particulate free. An isotonic formulation is in some embodiments used. Generally, additives for isotonicity may include sodium chloride, dextrose, mannitol, sorbitol and lactose. In some cases, isotonic solutions such as phosphateVU Ref.24027 / MBF Ref.093386-0031-WO01 buffered saline can be used. Stabilizers include gelatin and albumin. In some embodiments, a secondary treatment for preterm labor is added to the composition such as, but not limited to, nifedipine, indomethacin, magnesium sulfate, terbutaline, ritodrine, atosiban, barusiban, nitroglycerin, makena, hydroxyprogesterone caproate, and a progesterone supplement. In some embodiments, anti-inflammatories, immunosuppressants, antibiotics, analgesics, iron supplements, calcium supplements, vitamin D supplements, or combinations thereof are added to the composition.

[0053] The composition may further comprise a pharmaceutically acceptable excipient. The pharmaceutically acceptable excipient may be functional molecules as vehicles, adjuvants, carriers, or diluents. The term “pharmaceutically acceptable carrier,” may be a non-toxic, inert solid, semi-solid or liquid filler, diluent, encapsulating material or formulation auxiliary of any type. Pharmaceutically acceptable carriers include, for example, diluents, lubricants, binders, disintegrants, colorants, flavors, sweeteners, antioxidants, preservatives, glidants, solvents, suspending agents, wetting agents, surfactants, emollients, propellants, humectants, powders, pH adjusting agents, and combinations thereof. The pharmaceutically acceptable excipient may be a transfection facilitating agent, which may include surface active agents, such as immune- stimulating complexes (ISCOMS), Freunds incomplete adjuvant, LPS analog including monophosphoryl lipid A, muramyl peptides, quinone analogs, vesicles such as squalene and squalene, hyaluronic acid, lipids, liposomes, calcium ions, viral proteins, polyanions, polycations, or nanoparticles, or other known transfection facilitating agents. The transfection facilitating agent may be a polyanion, polycation, including poly-L-glutamate (LGS), or lipid.

[0054] In another aspect, the present disclosure provides a pharmaceutical composition comprising a compound as described herein, or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier. In a further embodiment, the composition comprises at least two pharmaceutically acceptable carriers, such as those described herein. For purposes of the present disclosure, unless designated otherwise, solvates and hydrates are generally considered compositions. Preferably, pharmaceutically acceptable carriers are sterile. The pharmaceutical composition can be formulated for particular routes of administration such as oral administration, vaginal administration, nasal administration, parenteral administration, and rectal administration, etc. In addition, the pharmaceutical compositions of the present disclosure can be made up in a solid form (including without limitation capsules, tablets, pills, granules, powders or suppositories), in a liquid form (including without limitation solutions, suspensions or emulsions), or aerosolized. The pharmaceutical compositions can be subjectedVU Ref.24027 / MBF Ref.093386-0031-WO01 to conventional pharmaceutical operations such as sterilization and / or can contain conventional inert diluents, lubricating agents, or buffering agents, as well as adjuvants, such as preservatives, stabilizers, wetting agents, emulsifiers and buffers, etc. In some embodiments, the pharmaceutical compositions are tablets or gelatin capsules comprising the active ingredient together with one or more of: diluents, e.g., lactose, dextrose, sucrose, mannitol, sorbitol, cellulose and / or glycine^ lubricants, e.g., silica, talcum, stearic acid, its magnesium or calcium salt and / or polyethyleneglycol^ binders, e.g., magnesium aluminum silicate, starch paste, gelatin, tragacanth, methylcellulose, sodium carboxymethylcellulose and / or polyvinylpyrrolidone^ disintegrants, e.g., starches, agar, alginic acid or its sodium salt, or effervescent mixtures^ and absorbents, colorants, flavors and sweeteners.

[0055] Certain injectable compositions are aqueous isotonic solutions or suspensions, and suppositories are advantageously prepared from fatty emulsions or suspensions. Said compositions may be sterilized and / or contain adjuvants, such as preserving, stabilizing, wetting or emulsifying agents, solution promoters, salts for regulating the osmotic pressure and / or buffers. In addition, they may also contain other therapeutically valuable substances. Said compositions may be prepared according to conventional mixing, granulating or coating methods, respectively, and contain about 0.1-100%, or about 0.1-75%, or contain about 1-50%, of the active ingredient.

[0056] In some embodiments, the composition may comprise a therapeutically effective amount of aprepitant. In some embodiments, the composition may further comprise a pharmaceutically acceptable carrier.

[0057] In some embodiments, the composition may comprise a therapeutically effective amount of ciclesonide. In some embodiments, the composition may further comprise a pharmaceutically acceptable carrier. In some embodiments, the composition may comprise a therapeutically effective amount of ciclesonide and a therapeutically effective amount of atosiban. The ciclesonide and the atosiban may be at any ratio. In some embodiments, the ciclesonide and the atosiban may be at a ratio of from about 361:1 to about 0.09:1. In some embodiments, the ciclesonide and the atosiban may be at a ratio of about 11.8:1. In some embodiments, the composition may comprise a therapeutically effective amount of ciclesonide and a therapeutically effective amount of ivermectin. The ciclesonide and the ivermectin may be at any ratio. In some embodiments, the ciclesonide and the ivermectin may be at a ratio ofVU Ref.24027 / MBF Ref.093386-0031-WO01 from about 57:1 to about 0.01:1. In some embodiments, the ciclesonide and the ivermectin may be at a ratio of about 1.8:1.

[0058] In some embodiments, the composition may comprise a therapeutically effective amount of ivermectin. In some embodiments, the composition may further comprise a pharmaceutically acceptable carrier. In some embodiments, the composition may comprise a therapeutically effective amount of ivermectin and a therapeutically effective amount of indomethacin. The indomethacin and the ivermectin may be at any ratio. In some embodiments, the indomethacin and the ivermectin may be at a ratio of from about 122:1 to about 0.03:1. In some embodiments, the indomethacin and the ivermectin may be at a ratio of about 7.7:1.

[0059] In some embodiments, the composition may comprise a therapeutically effective amount of nelfinavir mesylate. In some embodiments, the composition may further comprise a pharmaceutically acceptable carrier. In some embodiments, the composition may comprise a therapeutically effective amount of nelfinavir mesylate and a therapeutically effective amount of indomethacin. The indomethacin and the nelfinavir mesylate may be at any ratio. In some embodiments, the indomethacin and the nelfinavir mesylate may be at a ratio of from about 50:1 to about 0.01:1. In some embodiments, the indomethacin and the nelfinavir mesylate may be at a ratio of about 3:1. 4. Administration

[0060] The compounds or pharmaceutically acceptable salts thereof as detailed herein, or at least one component thereof, may be administered or delivered to a subject or a cell of the subject. Compounds or pharmaceutically acceptable salts thereof as detailed herein, or at least one component thereof, may be administered or delivered to a subject or a cell of the subject in a therapeutically effective amount. Such compositions can be administered in dosages and by techniques well known to those skilled in the medical arts taking into consideration such factors as the age, sex, weight, and condition of the particular subject, and the route of administration. The presently disclosed compounds or pharmaceutically acceptable salts thereof, or compositions comprising the same, may be administered to a subject by different routes including orally, parenterally, sublingually, transdermally, rectally, transmucosally, topically, intranasal, intravaginal, via inhalation, via buccal administration, intrapleurally, intravenous, intraarterial, intraperitoneal, subcutaneous, intradermally, epidermally, intramuscular, intranasal,VU Ref.24027 / MBF Ref.093386-0031-WO01 intrathecal, intracranial, and intraarticular, or combinations thereof. In some embodiments, the composition as described herein may be administered orally, vaginally, rectally, subcutaneously, or by intramuscular injection.

[0061] The term “a therapeutically effective amount” of a compound as described herein refers to an amount of the compound as described herein that will elicit the biological or medical response of a subject, for example, reduction or inhibition of an enzyme or a protein activity, or ameliorate symptoms, alleviate conditions, slow or delay disease progression, or prevent a disease, etc. In one non-limiting embodiment, the term “a therapeutically effective amount” refers to the amount of a compound as described herein that, when administered to a subject, is effective to, at least partially alleviate, inhibit, prevent and / or ameliorate a condition or a disorder mediated by preterm labor or preterm birth.

[0062] In some embodiments, a composition described herein may be administered more than once. In some embodiments, a composition described herein may be administered to a subject more than once per day. In some embodiments, a composition described herein may be administered after the onset of preterm labor. In some embodiments, a composition described herein may be administered before the onset of preterm labor. In some embodiments, a composition described herein may be administered repeatedly beginning about the 18th to 22nd week of gestation until about the 37th week of gestation. In some embodiments, a composition described herein may be administered repeatedly for about 14 to 19 weeks.

[0063] In some embodiments, a composition described herein may be administered with a secondary treatment for preterm labor. The secondary treatment for preterm labor may be nifedipine, indomethacin, magnesium sulfate, terbutaline, Ritodrine, Atosiban, Barusiban, nitroglycerin, Makena, hydroxyprogesterone caproate, progesterone supplementation, or cervical cerclage. a. Cell Types

[0064] Any of the delivery methods and / or routes of administration detailed herein can be utilized with a myriad of cell types, for example, those cell types currently under investigation for preterm labor or preterm birth, including, but not limited to, uterine cells, uterine myometrial cells, myometrial cells, smooth muscle cells, sensory neurons, neurons of the autonomic nervous system, motor neurons, fibroblasts, epithelial cells, skeletal muscle progenitors, smoothVU Ref.24027 / MBF Ref.093386-0031-WO01 muscle progenitors, human skeletal myoblasts, or other myogenic progenitor cells. Cells can be modified in vivo or ex vivo. 5. Methods for Treating or Preventing Preterm Labor or Decreasing Preterm Birth

[0065] Provided herein are methods of treating or preventing preterm labor or decreasing preterm birth in a pregnant subject in need thereof. The methods may include administering to the subject a composition comprising a therapeutically effective amount of a compound described herein. The subject may exhibit one or more of the following before administration of a composition described herein: recurrent uterine contractions^ cervical dilation^ cervical effacement^ and a cervical change. The methods described herein may further comprise monitoring or confirming a reduction in effacement, uterine contractions, cervical dilation, or a combination thereof after the administration of the composition. The methods described herein may prevent the onset of preterm labor. The methods described herein may prolong gestation.

[0066] Aprepitant, its current prodrug Fosaprepitant, or a composition comprising aprepitant or any other prodrug thereof may be administered alone or concomitantly with ciclesonide, ivermectin, nelfinavir mesylate, atosiban, indomethacin, or compositions comprising the same, or combinations thereof, for treating or preventing preterm labor or decreasing preterm birth in a pregnant subject. Ciclesonide, any prodrug thereof, or a composition comprising ciclesonide or any prodrug thereof may be administered alone or concomitantly with aprepitant, ivermectin, nelfinavir mesylate, atosiban, indomethacin, or compositions comprising the same, or combinations thereof, for treating or preventing preterm labor or decreasing preterm birth in a pregnant subject. Ivermectin, any prodrug thereof, or a composition comprising ivermectin or any prodrug thereof may be administered alone or concomitantly with aprepitant, ciclesonide, nelfinavir mesylate, atosiban, indomethacin, or compositions comprising the same, or combinations thereof, for treating or preventing preterm labor or decreasing preterm birth in a pregnant subject. Nelfinavir mesylate, any prodrug thereof, or a composition comprising nelfinavir mesylate or any prodrug thereof may be administered alone or concomitantly with aprepitant, ciclesonide, ivermectin, atosiban, indomethacin, or compositions comprising the same, or combinations thereof, for treating or preventing preterm labor or decreasing preterm birth in a pregnant subject.VU Ref.24027 / MBF Ref.093386-0031-WO01

[0067] In some embodiments, aprepitant, its prodrug fosaprepitant, any other prdrug of aprepitant, or a composition comprising aprepitant or any prodrug thereof may be administered for treating or preventing preterm labor or decreasing preterm birth in a pregnant subject.

[0068] In some embodiments, ciclesonide, any prodrug thereof, or a composition comprising ciclesonide or any prodrug thereof may be administered for treating or preventing preterm labor or decreasing preterm birth in a pregnant subject. In some embodiments, ciclesonide and atosiban, or compositions comprising the same, may be administered for treating or preventing preterm labor or decreasing preterm birth in a pregnant subject. The ciclesonide may be administered before, at the same time, or after the atosiban. The ciclesonide and the atosiban may be administered to the subject at any ratio. In some embodiments, the ciclesonide and the atosiban may be administered to the subject at a ratio of from about 361:1 to about 0.09:1. In some embodiments, the ciclesonide and the atosiban may be administered to the subject at a ratio of about 11.8:1.

[0069] In some embodiments, ciclesonide and ivermectin, or compositions comprising the same, may be administered for treating or preventing preterm labor or decreasing preterm birth in a pregnant subject. The ciclesonide may be administered before, at the same time, or after the ivermectin. The ciclesonide and the ivermectin may be administered to the subject at any ratio. In some embodiments, the ciclesonide and the ivermectin may be administered to the subject at a ratio of from about 57:1 to about 0.01:1. In some embodiments, the ciclesonide and the ivermectin may be administered to the subject at a ratio of about 1.8:1.

[0070] In some embodiments, ivermectin and indomethacin, or compositions comprising the same, may be administered for treating or preventing preterm labor or decreasing preterm birth in a pregnant subject. The ivermectin may be administered before, at the same time, or after the indomethacin. The indomethacin and the ivermectin may be administered to the subject at any ratio. In some embodiments, the indomethacin and the ivermectin may be administered to the subject at a ratio of from about 122:1 to about 0.03:1. In some embodiments, the indomethacin and the ivermectin may be administered to the subject at a ratio of about 7.7:1.

[0071] In some embodiments, nelfinavir mesylate and indomethacin, or compositions comprising the same, may be administered for treating or preventing preterm labor or decreasing preterm birth in a pregnant subject. The nelfinavir mesylate may be administered before, at the same time, or after the indomethacin. The indomethacin and the nelfinavirVU Ref.24027 / MBF Ref.093386-0031-WO01 mesylate may be administered to the subject at any ratio. In some embodiments, the indomethacin and the nelfinavir mesylate may be administered to the subject at a ratio of from about 50:1 to about 0.01:1. In some embodiments, the indomethacin and the nelfinavir mesylate may be administered to the subject at a ratio of about 3:1. 6. Examples

[0072] The foregoing may be better understood by reference to the following examples, which are presented for purposes of illustration and are not intended to limit the scope of the invention. The present disclosure has multiple aspects and embodiments, illustrated by the appended non-limiting examples. Example 1 Materials and Methods

[0073] Primary Uterine Myometrial Cells. Human myometrial tissue biopsies were obtained at the time of cesarean section. Reproductive-age (18-45 years) women receiving a scheduled or repeat cesarean section were recruited, fully informed, and consented to the study. Inclusion criteria included: breech presentation, previous cesarean section, fetal anomaly, fetal distress, or placental previa. Exclusion criteria included: clinical or histological signs of vaginal / chorioamniotic / intrauterine inflammation / infection, currently taking vasopressors or bronchodilators, or insulin-controlled diabetics. Samples were placed into either sterile cold 1XHBSS or room-temperature 1XPBS, and then transported to the laboratory for myometrial cell isolation or ex vivo isometric contractility assays, respectively.

[0074] Primary myometrial cells were isolated, selectively enriched for smooth muscle cells, and cultured as previously described (Herington et al., PLoS One.2015^ 10(11): e0143243^ Siricilla et al., Pharmacol Res.2019^146:104268^ and Siricilla et al., Pharmacol Res.2023^ 195:106876). Primary myometrial cells were unpassaged and became near-confluent 10-15 days post-isolation, with media (DMEM supplemented with 10% fetal bovine serum (FBS), 25 mM HEPES, 100 U / mL penicillin-streptomycin) changed every 3 days. The cells were dissociated at near-confluency using 0.25% Trypsin-EDTA, then plated in black-walled 384-well plates (Greiner Bio-One, Kremsmünster, Austria) for high-throughput screening Ca2+- mobilization assays as previously described (Siricilla et al., Pharmacol Res.2019^146:104268).VU Ref.24027 / MBF Ref.093386-0031-WO01

[0075] High-throughput drug discovery assay. A dual-addition Ca2+-mobilization assay was performed exactly as previously described (Siricilla et al., Pharmacol Res.2019^146:104268). Briefly, Fluo-4AM (2 ^M) was loaded into cells (4,000 / well) in 384-well microtiter plates. A Panoptic (WaveFront Biosciences, Franklin, TN) integrated pipettor was used to add compounds during the first addition, add oxytocin (OT^ Sigma, Burlington, MA) during the second addition, as well as continuously measure fluorescence in real time. Control wells were used to calculate hit-compounds, and received 0.1% DMSO (vehicle) during the compound addition, followed by EC80 of OT which was determined by performing three-fold 13-point concentration response analysis of OT. Test wells received 10 ^M compounds from the SelleckChem (Houston, TX) collection of FDA-approved drugs.

[0076] Comparative concentration-dependent responses. Hit-compounds were assayed at three-fold 10-point titrations starting at 30 ^M in triplicate, as previously described to compare the Emaxand IC50 values between myometrial and vascular SMCs (Siricilla et al., Pharmacol Res.2019^146:104268). Oxytocin was utilized as an agonist during the second addition for myometrial cells, while U46619 (a thromboxane A2 receptor agonist^ Cayman Chemicals, Ann Arbor, MI) (Pfister, Prostaglandins Other Lipid Mediat., 2008^ 87: 54-61^ Lin et al., World J. Gastroenterol., 2005^ 11: 2306-2312) was used in aorta VSMCs. On each day of screening, the OT and U46619 EC80 values were calculated to determine the ability of test compounds to inhibit this submaximal concentration. Primary human aorta cells (ATCC® PCS-100-012™) cultured in complete media as previously described (Siricilla et al., Pharmacol Res. 2019^146:104268). All concentration response curves (CRCs) were performed in triplicate. GraphPad prism was used to obtain IC50s through non-linear analysis.

[0077] Combination high-throughput screen. An 8X8 dose matrix was used to evaluate the combination effects between two serially-diluted single-compound concentrations. The single- drug IC50 values listed in TABLE 4 were utilized, with three 2-fold titrations above and below the IC50 value. Control (no drug) additions were included in the dose matrix (FIG.4A, FIGS. 4E-F, and FIGS.4I-J). The first row and the first column of each matrix was the individual drug concentrations used to compare the effect of combination. The high-throughput Ca2+-assay was executed exactly as described above, and in 384-well format, allowing for 6 drug combinations per plate.

[0078] Precise titrations of drugs into a compound plate were performed using an Echo555 within the Vanderbilt HTS Facility. To account for position bias and edge effects, the matricesVU Ref.24027 / MBF Ref.093386-0031-WO01 were repeated at different positions of the 384-well plate. Also, the z-factor determined during the optimization of the Ca2+assay (Herington et al., PLoS One.2015^ 10(11): e0143243 and Siricilla et al., Pharmacol Res.2019^146:104268), ensures lack of position effects. Raw data was first analyzed using Waveguide, as described above and Siricilla et al., Pharmacol Res. 2019^146:104268. After calculating the percent response for each compound concentration examined, synergistic combinations were determined using Combenefit software that provided synergy scores and heat-maps to visualize models of Bliss independence, Highest single agent, and Loewe additivity. The same percent response was also analyzed using an in-house developed tool Multidimensional Synergy of Combinations (MuSyC) as a secondary confirmation (Meyer et al., Cell Syst.2019^ 8(2): 97-108). Synergistic combinations of hit- antagonists were compared to the results from combinations of current tocolytics: Atosiban, Indomethacin, and Nifedipine. After selecting up to 3 fixed ratios of synergistic drug combinations, a concentration-response analysis was performed as above, to confirm whether the synergy was a result of increased efficacy and / or potency. Promising synergistic concentration ratios of antagonist drugs were tested on vascular SMCs to confirm that they retain their uterine selectivity. Synergistic combinations of antagonist drugs that exhibit ^ 5-fold difference between myometrial and VSMCs were ranked by their lowest combined doses required to achieve IC50, as well as compared to the results from combinations of current tocolytics.

[0079] Cell viability assay. Cytotoxicity of drug combinations on myometrial (HTERT-HM), kidney (RPTEC), and liver (HEPG2) cells were assessed using a standard WST-1 cell viability assay. The HTERT-HM (Condon et al., Biol Reprod.2002^ 67(2): 506-14), RPTEC (ATCC® PCS-400-010™), and HEPG2 (ATCC® HB-8065™) cells were cultured in complete DMEM-F12 (Gibco 11-039-02, Thermo Fisher, Waltham, MA), RPTEC media (ATCC® PCS-400-030™), EMEM (ATCC® 30-2003™), respectively (DMEM-F12, RPTEC, and EMEM media all were supplemented with 10% FBS, 1% pencillin-streptomycin). RPTEC media was supplemented with triiodothyronine, rh EGF, hydrocortisone hemisuccinate, rh Insulin, epinephrine, L- Alanyl-L- Glutamine, transferrin^ ATCC® PCS-400-040™. The cells were transferred to 96-well cell culture plates (2 x 104cells per well) using the respective media. Serially diluted aliquots of each test combination at concentrations ranging from 0.78–200 ^M (a final volume of 100 ^L) were added to the cells. Control compounds with known toxicity such as napabucasin were included on each plate. The plates were incubated for 72 hours and at the end of incubation, 10 ^L of WST-1 (Roche) was added to each well. Absorbance at 450 nm and 600 nm were read 2VU Ref.24027 / MBF Ref.093386-0031-WO01 hours after incubation with WST-1 and percent reduction in cell viability was calculated. After deduction of background absorbance, ten-point concentration-response curves were generated for each combination. Non-linear regression analysis was performed and IC50 values were determined from the CRCs using GraphPad prism software.

[0080] Ex vivo myometrial contractility. An isometric contractility organ bath assay was performed as previously described (Herington et al., PLoS One.2015^ 10(11): e0143243^ Siricilla et al., Pharmacol Res.2019^146:104268^ Siricilla et al., Pharmacol Res.2023^ 195: 106876). Briefly, mouse and human myometrial strips (12 mm X 5 mm X 1 mm, N = 6–11 from 4 to 5 mice and about 8 humans) were submerged in a heated and oxygenated (37°C, 95% O2– 5% CO2) Radnoti LLC tissue bath containing Kreb’s Bicarbonate Solution. Tissue was placed under 3 g of tension, washed with Kreb’s Bicarbonate Solution, and treated with KCl, as previously described (Siricilla et al., Pharmacol Res.2023^ 195: 106876). Rhythmic contractions were typically established within about 2–3 h. Following the establishment of rhythmic, spontaneous contractions, cumulative concentrations (1 nM to 0.1 mM) of drugs or vehicle control were added to individual organ baths every 20 minutes. In a second set of experiments, cumulative concentrations (1 nM to 0.1 mM) of drug combinations at their synergistic fixed-ratios, as well as each of their single-drugs alone (at their respective concentrations reflective of the ratio) were added to individual organ baths. After the highest concentration of drug or vehicle examined, tissue viability at the end of the experiment was evaluated using exposure to 75 mM KCl.

[0081] Isometric contractions were recorded using PowerLab / 8 SP (ADInstruments, Dunedin 9016, New Zealand) equipment and analyzed with LabChart 7 Pro software (ADInstruments, Dunedin 9016, New Zealand). Contractile activity was assessed by AUC / duration (integral relative to baseline divided by the duration of each treatment period assessed). All treatment data were expressed as a percent change of baseline spontaneous contractile activity. Data were analyzed and expressed as mean ± SD. Non-linear regression analyses yielded CRCs for calculation of IC50 and Emax. Comparisons of fit determined whether the non-linear fit lines between the drugs were significantly different (p<0.05) from DMSO using GraphPad Prism Software.

[0082] Assessment of fetal DA constriction. CD-1 female and male mice were bred in a restricted 3 hr window (8–11 am), and the presence of a copulatory plug was considered day 1 of pregnancy. The vasoreactivity of term (gestation day 19) fetal mouse DAs (N=7–9,VU Ref.24027 / MBF Ref.093386-0031-WO01 representing at least three different litters) was determined using cannulated, pressurized vessel myography, and computer-assisted videomicroscopy, as previously described (Reese et al., Prostag Oth Lipid M.2009^ 88(3-4): 89-96 and Pfaltzgraff et al., J. Mol. Cell Cardiol.2014^ 69: 88-96. Briefly, the fetal DA was mounted in custom myography chambers placed on inverted microscopes equipped with a digital image capture system (IonOptix) to record changes in the intraluminal vessel diameter. The pressure was increased in 5-mmHg increments to 20 mmHg, followed by treatment with 50 mM KCl deoxygenated modified Krebs buffer to determine vessel viability and peak contractility. The vessels were then changed from a flow-through system to a 20 mL recirculating system and allowed to equilibrate for an additional 20 minutes. Baseline lumen diameter was recorded prior to adding cumulative concentrations (1 nM to 0.1 mM) of drugs or vehicle control every 20 minutes. After the highest concentration of drug or vehicle examined, vessel viability was determined via exposure to 50 mM KCl. All treatment data were expressed as a percent change in baseline lumen diameter.

[0083] In vivo therapeutic efficacy and safety in a mouse model of induced-preterm labor. Experiments involving mice were conducted in accordance with the regulations provided by the National Institutes of Health Guide for the Care and Use of Laboratory Animals, using protocols approved by the Institutional Animal Care and Use Committee at Vanderbilt University Medical Center. For in vivo mouse studies, individual mice were randomized to receive indicated treatment and to detect 80% power with a=0.05 based on prior data using the same model of induced-PTL. On gestation day 15, mice (N ^ 5 per group) were administered mifepristone (s.c.^ Sigma M8046) at the lowest dose (30 μg) required to induce PTB in 100% mice. Preterm birth was defined as delivery 24 hours (hrs) before the average normal term delivery. The initial starting dose for each drug was 30 mg / kg (equal to 10X Emax from ex vivo organ bath studies), given 5 hours post-MIF (after the onset of in vivo uterine contractions), followed by treatments every 12 hours until 7am on gestational day 18. Dosing strategies were optimized based on improvement in outcomes of interest to values comparable to untreated controls: maternal weight gain (gd15-delivery), term birth rates, litter size (# pups born alive, dead, resorbed), pup viability (# pups born alive at term relative to total litter size), pup weight, placental weight. Additional doses of aprepitant tested included 50 mg / kg and 70 mg / kg. The dose of ciclesonide was reduced to (7.5, 15, 30, 40, or 50 mg / kg) at 5 hours and 17 hours (i.e. gd16 am) post-MIF. In a second dosing strategy for ciclesonide, a third treatment was added on gd17 am. Subsequently, a fourth treatment was added on gd18 am as a 3rd dosing strategy. In the 4th and 5th dosing strategy, a loading dose (20 or 30 mg / kg) was given 5 hours post-MIF, followedVU Ref.24027 / MBF Ref.093386-0031-WO01 by once daily am injections of various loading doses (1.8, 3.8, 7.5, or 15 mg / kg). A dose of 30 mg / kg loading and 1.8 mg / kg maintenance was used for synergistic drug combinations containing ciclesonide with either atosiban (FR 11.8:1^ 2.54 mg / kg loading and 0.15 mg / kg maintenance) or ivermectin (FR 1.8:1^ 16.7 mg / kg loading and 1 mg / kg maintenance). A dose of 1.7 mg / kg given 5 hours post-MIF, followed by every 12 hours until gd18 morning was used for synergistic drug combinations containing indomethacin with either ivermectin (FR 7.7:1^ 0.218 mg / kg) or nelfinavir mesylate (FR 3:1^ 0.57 mg / kg). Vehicles included 0.21 mL / kg ethanol + 0.12 mL / kg DMSO for aprepitant and indomethacin, or 0.33 mL / kg ethanol for other test drugs and combinations, which were further dissolved in 150 μL sesame oil prior to s.c. treatment.

[0084] The onset of delivery was monitored using infrared video surveillance and defined as delivery of the first pup. Preterm birth was defined as delivery occurring 24 hrs before the average onset of term delivery of untreated mice. At least three pups per litter were used to assess skin maturation, as determined by the toluidine blue dye method. To do so, pups were euthanized under CO2, dehydrated by immersing in 100% methanol for 1-2 min, and rinsed with PBS. Pups were then immersed in 0.1% toluidine blue in PBS for 1 hr, rinsed with PBS, and photographed. At least three pup lungs per litter were formalin-fixed followed by routine histological processing and H&E staining at the Vanderbilt Translational Pathology Shared Resource. For morphometric analysis, brightfield images (N=6-10 per pup) of H&E-stained lung sections were captured using a Keyence BZ-X810 and 40x objective. Large airways and vessels were avoided when imaging sections. Lung morphometric analysis was performed in a blinded fashion regarding which samples correspond to treatment groups. Airspace volume density (ASVD) was determined by importing TIFF files into Napari and in-house built software (Alveoleye^ www.napari-hub.org / plugins / AlveolEye). Example 2 Identification Of 52 FDA-Approved Drugs as Hit-Antagonists of OT-Induced Ca2+-Release

[0085] In an effort to repurpose drugs for tocolytic use, a systematic screening strategy was used to probe the potential of FDA-approved drugs for novel tocolytic use (FIG.1A). A library of 1,180 FDA-approved drugs were screened for their ability to inhibit oxytocin (OT)-induced calcium mobilization from human primary myometrial cells isolated from tissue obtained at the time of cesarean delivery at Vanderbilt University Medical Center (VUMC). Subsequently, the repurposing potential of these drugs for novel tocolytic use were probed by examining the: (i) inVU Ref.24027 / MBF Ref.093386-0031-WO01 vitro uterine-selectivity by comparing concentration-response between myometrial cells and a major maternal off-target site, vascular smooth muscle cells (VSMCs), (ii) synergy in combination with one another, as well as with current clinical tocolytics (Atosiban, Nifedipine, Indomethacin, and Terbutaline), (iii) cytotoxic effects, (iv) ex vivo efficacy, potency and tissue- selectivity between myometrial contractility and constriction of ductus arteriosus (DA), a major fetal off-target of current tocolytics, and (v) in vivo efficacy to delay delivery in a mouse model of induced preterm labor leading to preterm birth and impact on neonatal outcome. Each of the above screening steps narrowed hit-drugs into leads based on strict criteria of uterine selectivity, favorable therapeutic index (TI), tissue level confirmation assays and preclinical in vivo testing in mouse models.

[0086] Given that an increase in intracellular Ca2+-levels is the final common pathway controlling uterine myometrial contractions for labor, and oxytocin (OT) is the most potent endogenous uterine contractile agonist, an in vitro high-throughput drug discovery assay was used to screen the 1,180 small molecule drugs within the SelleckChem FDA library for antagonists of OT-induced intracellular Ca2+-release (FIG.1B).

[0087] A total of 52 drugs were found to inhibit myometrial calcium-mobilization (FIG.1C and TABLE 1) from varying drug classes (FIG.1D), including: antifungal agents, anti-asthmatic agents, anti-parasitic, and antiemetics. Of these, 13 drugs were more potent than the positive control drug atosiban - an oxytocin receptor antagonist and current tocolytic with European Approval. TABLE 1. Hit-antagonists of OT-induced Ca2+-mobilization identified from SelleckChem FDA library of 1180 drugs. Hit-antagonists %Anidulafungin Crystal violet Evacetrapib Bexarotene Cetylpyridinium Chloride Ivermectin Conivaptan HCl Ivacaftor (VX-770) Benzethonium chloride Ospemifene Cetrimonium Bromide BenzbromaroneVU Ref.24027 / MBF Ref.093386-0031-WO01VU Ref.24027 / MBF Ref.093386-0031-WO01 Indomethacin10 Bolded values represent drugs that were more potent than atosiban in inhibiting Ca2+mobilization frommyometrial cells. Example 3 Prioritization of Hit-Drugs Based on Efficacy, Potency, and Myometrial-Selectivity

[0088] The primary limitation of current therapeutics used to arrest PTL, such as Nifedipine and Indomethacin, is their off-target effects on maternal vascular smooth muscle cells (VSMCs) and fetal ductus arteriosus smooth muscle cells (SMCs). Given that adult primary aorta VSMCs displayed the most similarity to primary human myometrial cells when comparing their transcriptome profile, this specific VSMC was selected for counter-screening efforts. To test hit- antagonists for uterine-selectivity, U46619 (an agonist of the thromboxane A2 receptor) was used, instead of OT, to induce intracellular Ca2+-release from VSMCs since it is established as the most universal potent agonist of SMCs from any vascular bed. Thus, by using the Ca2+- mobilization assay and primary human aorta vascular smooth muscle cells, whether hit-drugs were uterine-selective based on their lack of activity in vascular SMCs or effect on Ca2+- mobilization in myometrial SMCs at ^ 5-fold activity compared to VSMCs was examined. Using this criterion, 12 hit-antagonists that are myometrial selective were identified (FIGS.1E-L and TABLE 2). Fosaprepitant Dimeglumine and ciclesonide were found to be the most selective at an Emax259x and 20x greater in myometrial cells compared to that of aorta cells. Collectively, these results indicate that small molecule regulators that are selectively modulatory of uterine myometrial Ca2+-mobilization can be identified. TABLE 2. Summary of myometrial selective hit-antagonists of Ca2+-mobilization fromVU Ref.24027 / MBF Ref.093386-0031-WO01Reprotox database. (í) Drug did not have any inhibitory effect on Ca2+mobilization in vascular cells, therfore, no measurable IC50. To calculate fold-change, (-) was converted to >30uM. Example 4 Functional Classifications of Hit-Antagonist Drugs

[0089] To gain insight into the mechanism of action of the hit-drugs from the FDA library, a combination of the Pubchem (pubchem.ncbi.nlm.nih.gov) and DrugBank (drugbank.ca) databases was used, as well as the Integrated database for ADMET and adverse effect predictive Modeling (idaapm.helsinki.fi). The myometrial-selective drugs were found to be from diverse drug classes with different targets (FIG.1D, TABLE 2), which is ideal for exploration of drug combinations to increase efficacy while reducing off-target side effects. The Reprotox database (reprotox.org) was searched for all drugs listed in TABLE 2, and it was found that some were reported to increase the risk of congenital malformations in humans. For example: 1) Afatinib in animals has shown maternal toxicity, reduced fetal weights, and increased incidents of visceral and dermal variations. Also, Afatinib is avoided in pregnancy based on its mechanism of action, as it is expected to cause fetal harm when administered during pregnancy^ 2) use of Tazarotene is avoided in human pregnancy as fetal malformations, toxicity, and developmental and / or behavioral delays occurred when used during pregnancy. Example 5 Elimination of Cytotoxic CombinationsVU Ref.24027 / MBF Ref.093386-0031-WO01

[0090] To prioritize and eliminate compounds that are toxic to the target organ (uterus), and the major metabolic and excretory organs (liver and kidney), a WST-1 cell viability assay was used to assess the cytotoxicity of 10 uterine-selective compounds on myometrial (HTERT-HM), kidney (RPTEC), and liver (HEPG2) cells. An in vitro therapeutic index (TI) obtained by measuring a compound’s cytotoxicity at its given efficacious concentrations is commonly used to quantify the extent of a drug’s safety. Thus, the in vitro TI was calculated as the ratio of IC50 values from cell viability and Ca2+assays. Two single drugs (Fosaprepitant Dimeglumine and Ciclesonide) had a TI>5, which was considered favorable (TABLE 3). Crystal violet, napabucasin, and penfluridol exhibited toxicity <10 μM for all cell types examined and were therefore removed from further study. TABLE 3. In vitro therapeutic index of uterine selective, FDA-approved drugs.TI: Ratio of IC50from cell viability assay to IC50from Ca2+assay. Bolded values represent drug combinations that have a TI ^5. Example 6 Detection of Synergistic Drug CombinationsVU Ref.24027 / MBF Ref.093386-0031-WO01

[0091] Combination therapy allows greater drug efficacy or potency due to additive or synergistic effects, thereby reducing the toxicity or side effects associated with higher doses of single drugs. Thus, it was determined whether the in vitro TI of uterine-selective hit-antagonists could become favorable (>5) as a result of combination drug synergy. Applicants’ established high-throughput drug combination screen was used to identify synergistic fixed-ratios of 45 non- redundant combinations of 7 uterine-selective hit-antagonists and 3 current tocolytics (atosiban, indomethacin, and nifedipine) that use different mechanisms of action. These drug combinations were tested at 2-fold titrations above and below each drug’s IC50 value using an 8x8 dose matrix (FIG.2B), then analyzed for synergistic inhibition of intracellular Ca2+-release using Combenefit. The combenefit analyzed the % response data using three popular classic models, Bliss independence, Loewe additivity, and Highest Single Agent (HSA) to define drug interaction. Combenefit enables model-based quantification of drug combinations and provides a synergy score (Di Veroli et al., Bioinformatics.2016^ 32(18): 2866-8). While each of the above models have distinct advantages and disadvantages, a recent publication by Gilvary et al. found that Bliss independence corresponds best to clinical success of combinations within a screen of antineoplastic compounds (Gilvary et al., Trends Pharmacol Sci.2019^ 40(8): 555-564 and Vlot et al., Drug Discov Today.2019^ 24(12): 2286-2298). The Bliss independent model was found to be the strictest among other models to pick synergistic combinations.

[0092] Representative synergistic combinations of hit-drugs from the SelleckChem collection with each other or current tocolytic drugs are shown in FIG.2D-G. Bliss independence heat maps demonstrate the dose matrix with synergy or antagonism.

[0093] For further confirmation of synergies, concentration-response analysis of fixed-ratios of selected synergistic combinations was performed. Combinations that contained crystal violet and / or penfluridol that exhibited synergy were excluded from this step since they were determined as cytotoxic combinations as described in the following paragraph. 25 synergistic combinations that remained after this exclusion and after selecting up to 3 fixed ratios (at least 1, up to 3) of each synergistic drug combination were focused on. A CRC analysis was performed to confirm whether the synergy was a result of increased efficacy and / or potency. It was found that 15 / 25 (60%) synergistic drug combinations from Bliss resulted in increased efficacy, while 11 / 25 (44%) resulted in increased potency. Representative CRC curves for fixed-ratios of combinations are shown in FIG.2H-K. TABLE 4 shows the Emaxand IC50s obtained from the CRCs of fixed-ratios for each drug combination in comparison to the individual drugs and the identified synergistic combinations in efficacy and / or potency.VU Ref.24027 / MBF Ref.093386-0031-WO01 TABLE 4. Synergistic combinations of uterine selective, FDA-approved drugs with each other and with known tocolytics.Bolded values represent drug combinations that are synergistic compared to individual drugs in terms of the Emaxand IC50. *Concentrations of each drug in the combination at 50% inhibition of Ca2+-mobilization.

[0094] The following synergistic fixed ratios were confirmed with in vitro concentration response studies, based on either efficacy and / or potency (TABLE 5):VU Ref.24027 / MBF Ref.093386-0031-WO01 TABLE 5. Synergistic Fixed Ratios of FDA-Approved Drugs in vitro.Example 7 Elimination of Cytotoxic Combinations

[0095] To prioritize and eliminate the synergistic combinations that are toxic to target organ (uterus), and the major metabolic and excretory organs (liver and kidney), selected fixed-ratios of the combinations (total 30 fixed ratios) confirmed as synergistic through CRCs were subjected to cell viability assay to explore their cytotoxicity in these early stages of drugVU Ref.24027 / MBF Ref.093386-0031-WO01 discovery. To do so, WST-1 cell viability assays were performed to assess cytotoxicity on myometrial (HTERT-HM), kidney (RPTEC), and liver (HEPG2) cells. A therapeutic index (TI, ratio of IC50 values from cell viability and Ca2+assays) >5 was considered favorable.

[0096] TABLE 6 shows the safety index of all the synergistic combinations at different fixed- ratios. A total of 5 synergistic combinations of FDA-approved drugs had a favorable TI. TABLE 6. Therapeutic index of combinations of uterine selective, FDA-approved drugs.Therapeutic index (TI) equals the ratio of IC50 from cell viability assay to IC50 from Ca2+assay. Bolded values represent drug combinations that have a TI ^ 5. Concentration of drug A (left to the +) and drug B (right to the +) needed in combination to inhibit the OT induced Ca2+mobilization (or) cell viability by 50% (IC50). Example 8 Ex Vivo Tocolytic Efficacy, Potency, and Lack of Effect on Fetal DA Vasoreactivity

[0097] Next, ex vivo uterine myometrial (“UT myo”) contractility of the uterine selective drugs and the four synergistic combinations mentioned above (Ciclesonide + Atosiban, Ciclesonide + Ivermectin, Indomethacin + Ivermectin, and Indomethacin + Nelfinavir Mesylate) that inhibited Ca2+release in UT myo cells were examined. A well-established ex vivo isometric contractility assay was used as a secondary screen to examine the effect of hit-antagonist compounds on frequency, amplitude, and area under the curve (AUC^ Herington et al., PLoS One.2015^ 10(11): e0143243^ Norman et al., Journal of Reproduction and Fertility.1997^VU Ref.24027 / MBF Ref.093386-0031-WO01 110(2): 249–54^ Oger et al., Biology of Reproduction.2004^70(2): 458–64^ Stymiest et al., The Journal of Organic Chemistry.2005^ 70(20): 7799–809^ Baumbach et al., Am J Obstet Gynecol. 2012^ 206(3): 254 e1-5^ Robinson et al., PLoS One.2012^ 7(9): e46278^ Crankshaw et al., Reproductive Sciences.2013^ 20(8): 882–90). Spontaneous myometrial contractions were recorded (measured in grams, g, of tension) for 20 minutes, followed by the addition of cumulative doses of vehicle (FIG.4A). Ciclesonide and fosaprepitant dimeglumine (the prodrug for aprepitant) significantly inhibited the contractile activity, relative to the vehicle control, at the highest three concentrations examined. Each single agent displayed ex vivo tocolytic efficacy >50% with potencies ranging from 0.6-1.3 ^M (FIGS.4C-J).

[0098] The following ex vivo tocolytic efficacies were observed for each synergistic fixed ratio compared to each single-drug at their respective concentration in the fixed ratio (FR): Ciclesonide + Atosiban at a ratio of 11.8:1 yielded 73% (0.2 ^M potency) whereas Ciclesonide alone yielded 57% (0.6 ^M potency) and Atosiban alone yielded 24% (0.8 ^M potency)^ Ciclesonide + Ivermectin at a ratio of 1.8:1 yielded 73% whereas Ciclesonide alone yielded 57% and Ivermectin alone yielded 42%^ Ivermectin + Indomethacin at a ratio of 1:7.7 yielded 85% (6.6 ^M potency) whereas Ivermectin alone yielded 46% (18 ^M) and Indomethacin alone yielded 62% (12 ^M potency)^ Nelfinavir Mesylate (NM) + Indomethacin at a ratio of 1:3 yielded 85% (0.6 ^M potency) whereas NM alone yielded 60% (0.9 ^M) and Indomethacin alone yielded 62% (12 ^M potency).

[0099] Undesirable off-target effects of a current tocolytic (Indomethacin) on in utero constriction of fetal DA is the major limitation of the drug for use as a tocolytic beyond 32 gestational weeks. This prompted the examination of the lead hit-antagonists for potential vasoconstrictive effects using cannulated DA from term gestation (day 19) mouse fetuses in an ex vivo pressure myography assay. Importantly, only nebivolol resulted in a significant decrease (constriction) of lumen diameter relative to vehicle-treated controls (FIGS.4C-J). Only nebivolol resulted in a significant decrease (constriction) of lumen diameter in ex vivo myography experiments using cannulated DA from term (day 19) fetuses. [000100] Among the combinations tested, Indomethacin in combination with Ivermectin (7.7:1) and Nelfinavie Mesylate each exhibited the greatest tocolytic efficacy (Emax=85%, FIGS.5C-D), while Ciclesonide and Atosiban (1:11.8) had the greatest potency (IC50=0.2 ^M^ FIG.5A). Example 9VU Ref.24027 / MBF Ref.093386-0031-WO01 In Vivo Therapeutic Efficacy to Delay Delivery and Improve Neonatal Outcomes in a Mouse Model of PTB [000101] The mifepristone (MIF)-induced mouse model of preterm labor (PTL) leading to preterm birth (PTB) was used to determine the pre-clinical efficacy of lead single (aprepitant and ciclesonide) and combination of drugs (ciclesonide + atosiban, ciclesonide + ivermectin, indomethacin + ivermectin, indomethacin + nelfinavir mesylate) to delay the timing of delivery and improve neonatal outcomes. As shown in FIGS.6A-D and FIGS.7A-D, MIF was administered on gestation day 15, followed by treatment with either vehicle, lead single or combination of drugs. The dosing strategies were optimized based on improvement in outcomes of interest to values comparable to untreated controls: maternal weight gain (gd15- delivery), term birth rates, litter size (# pups born alive, dead, resorbed), pup viability (# pups born alive at term relative to total litter size), pup weight, placental weight. Pups born at term to dams receiving lead single or combination of drugs after MIF-induction PTL were examined for their developmental maturation. Two tissues (skin and lungs) were used to determine developmental status (McCarthy et al., Biol Reprod.2018^ 99(5): 922-937) in comparison to the premature or mature developmental status of pups born from vehicle-treated MIF mice and untreated controls, respectively. [000102] Dose-response analysis of Aprepitant (FIG.6A) in vivo revealed that 50 mg / kg every 12 hours, subcutaneous injection (s.c.) was able to delay the timing of delivery until term (4.5 days after induction of PTL) in 50% of mice compared to vehicle treated mice that delivered within 24 hours after induction of PTL using Mifepristone. Other than term birth rate, 50 mg / kg aprepitant did not significantly impact any outcomes of interest (TABLE 7) relative to untreated term mice. The skin of pups born to aprepitant treated dams was impermeable to Toulidine blue dye, unlike the permeable skin of preterm gd16 pups born from vehicle-treated MIF-induced PTL dams (FIG.6C). Also, the airspace volume density (ASVD) of lungs collected at term birth pups born from aprepitant treated dams was statistically (p<0.0001) greater than preterm gd16 pups, but similar to those born from untreated dams (FIG.6D). These analyses indicate that the skin and lungs of pups born from dams receiving aprepitant to manage PTL are born developmentally mature. TABLE 7. In vivo efficacy of FDA-approved drugs in mouse models of PTB and neonatal outcomes.Placental weight (g)0.10 ± 0.00 -0.10 ± 0.00- -Maternal weight gain from gd15-delivery. Outcomes collected from term deliveries. (-) denotes data not applicable due to lack of term deliveries. (- -) denotes data not collected due to deliveries occurring overnight. Bolded values are significantly (p^0.05) compared to untreated (No Tx) controls. (*) denotes optimal drug dose. [000103] Dose-response analysis of Ciclesonide (FIG.7A) in vivo revealed that 30 mg / kg loading, 1.8 mg / kg maintenance, s.c.) was able to delay the timing of delivery until term (4.5 days after induction of PTL) in 67% of mice compared to vehicle treated mice that delivered within 24 hours after induction of PTL using Mifepristone. It was found that 30 mg / kg loading and 1.8 mg / kg maintenance doses of Ciclesonide resulted in significantly improved (p^0.05) maternal weight gain and increased (p^0.05) placental weights, with all other outcomes of interest statistically similar to untreated term dams and pups (TABLE 8). The skin of pups born to Ciclesonide treated dams was impermeable to Toulidine blue dye, unlike the permeable skin of preterm gd16 pups born from vehicle-treated MIF-induced PTL dams (FIG.7C). Also, the airspace volume density (ASVD) of lungs collected at term birth pups born from Ciclesonide treated dams was statistically (p<0.0001) greater than preterm gd16 pups, but similar to those born from untreated dams (FIG.7D). These analyses indicate that the skin and lungs of pups born from dams receiving ciclesonide to manage PTL are born developmentally mature. TABLE 8. In vivo efficacy of FDA-approved drugs in mouse models of PTB and neonatal outcomes. Ciclesonide No Tx DS #1 DS #2 DS #3 DS #4 DS #5 30 30 7.5 15 30 40 50 2030 20 30 30 L + 30 L + 30 L + L+L + L + L + 3.8 M 7.5 M 15 MVU Ref.24027 / MBF Ref.093386-0031-WO01[ 1 o t m w p dCiclesonide combinations were impermeable to Toulidine blue dye and had lung ASVD that was statistically similar to pups born from untreated dams (FIGS.8A-D). TABLE 9. In vivo efficacy and neonatal outcomes of ciclesonide synergistic combinations in a mouse model of PTB. No Tx CicleAtosC + A Cicle Iver C + IMaternal weight 12.0 ± gain (g)2.5 5.1 ± 1.7 -5.8 ± 2.110.5 ± 5.1 ± 1.7 1.74.9 ± 3.3Term birth rate10067 08067 67100Litter size (n)10 ± 110 ± 3 -12 ± 210 ± 3 10 ± 39 ± 4Pup viability (%) 99 ± 3 88 ± 17 - 98 ± 5* 77 ± 35 100 ± 0 97 ± 6 1.50 ± 1.38 ± 1.31 ± 1.38 ± 1.70 ± 1.40 ± Pup weight (g) 0.09 0.14 - 0.12* 0.14 0.17 0.16 Placental weight 0.10 ± 0.14 ± 0.10 ± 0.14 ± 0.11 ± 0.11 ± (g) 0.00 0.02 - 0.01* 0.02 0.04 0.03* Maternal weight gain from gd15-delivery. Outcomes collected from term deliveries. (-) denotes data not applicable due to lack of term deliveries. Values that are bolded or with* indicate p^0.05 compared to no-tx or Ciclesonide-alone, respectively.VU Ref.24027 / MBF Ref.093386-0031-WO01 [000105] The indomethacin + ivermectin (FR 7.7 :1) combination (1.7 mg / kg indomethacin + 0.22 mg / kg ivermectin) permitted term delivery for 67% treated dams, compared to 33% of dams treated with indomethacin alone and 50% of dams treated with ivermectin alone (FIG. 9B). All outcomes examined were statistically similar to those yielded from untreated dams (TABLE 10). The indomethacin + nelfinavir mesylate (FR 3:1) combination (1.7 mg / kg indomethacin + 0.57 mg / kg nelfinavir mesylate) permitted term delivery for 50% treated dams versus 33% of dams treated with indomethacin alone and 0% of dams treated with ivermectin alone (FIG.9B). All outcomes examined except pup weight were statistically similar to those yielded from untreated dams (TABLE 10). Moreover, pups born to either indomethacin combination had comparable DA patency at 4 hrs of life, skin that was impermeable to Toulidine blue dye, and lung ASVD that was similar to pups born at term from untreated dams (FIGS.9C- D). Given that acute or chronic indomethacin exposure can result in premature closure of thegain (g) 2.5 3.8 3.4 2.1 2.9 3.4 - 3.5 Term birth rate100 10033 506733 050Litter size (n)10 ± 1 10 ± 19 ± 1 13 ± 38 ± 310 ± 1 -9 ± 4Pup viability (%) 99 ± 3 100 ± 6 100 ± 0 100 ± 0 100 ± 0 100 ± 0 - 98 ± 4 1.50 ± 1.40 ± 1.54 ± 1.49 ± 1.51 ± 1.54 ± 1.41 ± Pup weight (g) 0.09 0.14 0.02 0.24 0.25 0.02 - 0.15* Placental weight 0.10 ± 0.10 ± 0.10 ± 0.10 ± 0.10 ± 0.10 ± 0.11 ± (g) 0.00 0.00 0.0 0.0 0.0 0.0 - 0.02 Maternal weight gain from gd15-delivery. Outcomes collected from term deliveries. (-) denotes data not applicable due to lack of term deliveries. Values that are bolded or with* indicate p^0.05 compared to no-tx or indomethacin-alone, respectively. [000106] These findings suggest that Aprepitant and Ciclesonide have practical application as promising single-agent tocolytics, and Ciclesonide + Atosiban (fixed ratio of 11.8:1), Ciclesonide + Ivermectin (fixed ratio of 1.8:1), and potentially Ivermectin + Indomethacin and NM +VU Ref.24027 / MBF Ref.093386-0031-WO01 Indomethacin, have practical application as combination tocolytics for the long-term management of PTL, allowing delivery of healthy offspring at term. [000107] Drug discovery efforts toward identification of novel tocolytics is vital considering the lack of safe, FDA-approved drugs used to treat preterm labor. Drug repurposing has the ability to bring a treatment to the clinic faster than the traditional approach. In fact, it is evident from the use of Nifedipine and Indomethacin for clinical management of PTL, although not FDA approved for PTL therapy that drugs approved for other indications can be repurposed to treatin vitro therapeutic index, ex vivo tocolytic efficacy, and ability to prolong gestation until term and improve neonatal outcomes in the mifepristone-induced mouse model of PTB. Furthermore, four synergistic combinations of the uterine-selective drugs were identified using a similar systematic screening approach, with their synergy confirmed in ex vivo and in vivo studies. [000108] The systematic screening strategies herein demonstrated numerous strengths to yield single and synergistic drug combinations. A phenotypic high-throughput screen allowed for identification of drugs from various classes. An in vitro counter-screen against vascular SMCs successfully removed hit-antagonists belonging to cardiovascular and antiasthmatic drug classes, allowing us to identify selective modulators of uterine smooth muscle. Fenoterol and Sasapyrine were identified as hit-antagonists of myometrial Ca2+-mobilization that lacked uterine selectivity due to their effect on aorta vascular smooth muscle cells. Fenoterol is an adrenergic beta-2 agonist that failed clinical trials as tocolytic due to its severe maternal cardiovascular effects. Sasapyrine is a non-steroidal anti-inflammatory drug that is warned against during late pregnancy due to concerns about premature ductal constriction and neonatal bleeding. [000109] The HTS combination screen was an important tool to rapidly and reliably identify synergistic drug combinations that inhibit in vitro and ex vivo human myometrial contractility withVU Ref.24027 / MBF Ref.093386-0031-WO01 improved potency and efficacy. Combination therapy is a promising approach to develop treatments for a wide range of disorders. Herein is the first reported large-scale combination screen performed in order to discover tocolytic combination therapy. There have been some reports on small scale studies combining current tocolytics (Atosiban, Nifedipine, Indomethacin), however, the reports are limited to combining these drugs at fixed concentrations and have failed to find meaningful synergy. Trial and error were relied on in the reports with regard to doses with no overarching rules to guide development. The study herein has taken the dose matrix approach which includes a wide range of concentrations, that helps to visualize complex drug interactions. The goals from this combination screen were two-fold. First, to determine if any useful synergy in potency, i.e., can the dose of each drug be reduced, thereby preventing off-target side effects, and second was to determine synergy in efficacy so as to improve the effect from either one drug alone. For example, Atosiban is not clinically successful due to its lack of efficacy in preventing PTL over placebo even after proven to be non-toxic at high concentrations. Herein, an effective combination of an FDA-approved drug with Atosiban was found to provide an overall improved effect towards PTL. As shown in herein, such combinations show improved potency and efficacy. The Atosiban and Indomethacin combination has synergy when combined at a ratio of 1:8, showing a reduced IC50 of Indomethacin allowing a reduced dose of drug to achieve a better effect. Four combinations with the highest probability of success (Ciclesonide+Atosiban, Ciclesonide+Ivermectin, Indomethacin+Ivermectin, Indomethacin+Nelfinavir Mesylate) were advanced to be tested ex vivo to determine their effect to inhibit contractility of human myometrial tissue. Using an ex vivo contractility assay, the tocolytic efficacy of promising hit-antagonists in mono- and combination therapy were examined prior to exploring their in vivo effect on timing of delivery and maternal / fetal health. [000110] To ensure success of a large-scale combination screen, in addition to choosing the correct experimental design, equally significant is choosing the right model to analyze the results and classify synergy, no effect or antagonism. Combenefit, a tool that quickly analyzes large data sets to provide synergy scores in three different popular models (Bliss, Loewe, and HSA) was used. Since many experimental and analytical factors could influence the observed synergy scores, comparing with different models are useful. Furthermore, MuSyC provides a clear separation between synergy in potency and synergy in efficacy through beta and alpha values. Synergistic combinations from both Bliss and MuSyc were included for the secondaryVU Ref.24027 / MBF Ref.093386-0031-WO01 confirmation through determining the CRC of selected fixed ratios to compare IC50s and Emaxof combinations with individual drugs. [000111] It has become increasingly important to identify and reduce risk early to improve the success of drug development. When drugs are approved by FDA for an indication, it has gone through a rigorous process to demonstrate its lack of toxicity at the therapeutic doses. However, when two drugs are combined, it is significant to rule out unintended synergistic toxicity at the synergistic concentrations early in the discovery process. In vitro cell-based toxicity assays can be used to rapidly assess potential safety issues. In vitro safety index is commonly used to quantify the extend of the safety at the desired efficacious conditions of a test drug combination. By including this step in the discovery approach here, potentially unsafe combinations were eliminated regardless of their synergistic efficacy / potency. The determination of each single- and combination drug’s in vitro TI, potentially unsafe drugs were able to be eliminated regardless of their efficacy, potency, or synergy. This included anti-cancer drugs and those with reported reproductive toxicity. [000112] The single drugs with favorable in vitro TI identified from this study include: 1) fosaprepitant dimeglumine, a prodrug of aprepitant that is a selective high-affinity antagonist of human neurokinin 1 (NK1) receptor and FDA-approved as an antiemetic, and 2) ciclesonide, a glucocorticoid approved for allergic rhinitis. The mechanisms of their novel tocolytic action are unknown. However, mouse and human studies have established that tachykinins and glucocorticoids are involved in the regulation of uterine smooth muscle contractility. Further support for investigation of Ciclesonide for treatment of PTL is provided by preclinical studies reporting this drug as a promising candidate to treat or prevent brochopulmonary dysplasia in neonates without the adverse neurodevelopmental effects observed with other synthetic glucocorticoids. Although the present study herein identified ivermectin as a hit-antagonist with an unfavorable in vitro TI, support for further investigation into its potential therapeutic benefit for PTL is supported by its potent ability to inhibit ex vivo uterine contractility (FIGS.4A-J) and in vivo efficacy at 0.22 mg / kg dose to allow 50% of dams to delivery at term with unhindered maternal and fetal outcomes examined. Moreover, ivermectin has established anti- inflammatory effects and a lack of reported neonatal deaths, maternal morbidity, preterm birth or low birthweight in association with its use during pregnancy. Example 10VU Ref.24027 / MBF Ref.093386-0031-WO01 Preclinical In Vivo Therapeutic Testing in Two Additional Mouse Models of Induced Preterm Birth [000113] The causes of PTB are multifactorial, yet in about 70% of cases, PTB follows spontaneous preterm labor (sPL) without a known maternal, fetal, or placental condition. Approximately half of the cases of idiopathic preterm labor are thought to be due to an overt or subclinical inflammatory response in fetal and maternal tissues. The current view is that most cases of spontaneous preterm labor with intra-amniotic inflammation occur in the absence of microorganisms, thus, representing a sterile inflammatory process. Women with spontaneous preterm labor and sterile intra-amniotic inflammation harbor an alarmin-enriched cytokine network containing interleukin-1 (alpha and beta), S100B, high-mobility group box-1, and others. Animal models have confirmed the implication of these alarmins to induce preterm birth. [000114] Currently, as shown herein, the in vivo efficacy of Aprepitant and Ciclesonide as single-agents, and 4 drug combinations (Ciclesonide+Atosiban, 1:11.8^ Ciclesonide+Ivermectin, 1:1.8^ Indomethacin+Nelfinavir Mesylate, 1:3^ and Indomethacin+Ivermectin, 1:7.7) to delay the timing of delivery and improve neonatal outcomes in the mifepristone-induced mouse model of preterm birth. Given the multiple etiologies for preterm labor in women, the in vivo efficacy of these single- and combination-drugs will be tested in two additional preclinical mouse models of induced-PTB using IL-1ȕ and S100B. [000115] To do so, CD-1 female and male mice will be bred in a restricted 3-hour window (8– 11 am), and the presence of a copulatory plug will be considered day 1 of pregnancy. On day 15 of pregnancy, mice (N ^ 5 per group) will be administered (s.c.) various doses of IL-1ȕ or S100B (1.25, 2.5, 5, 10 ^g) to establish the lowest dose required to induce PTB in 100% mice (prior to day 18.5 of pregnancy, which is 24 hours prior to normal term delivery). Based on the results of these studies, day 15 pregnant mice (average weight: 44.97 + 4.35 g) will be administered IL-1ȕ or S100B, followed by administration (s.c.) of treatment 2 hours after the onset of in vivo uterine contractions. Treatments will include either vehicle (0.7 mL / kg ethanol in 150 μL sesame oil, negative control), 5 mg / kg Nifedipine (positive control), Indomethacin alone (6.5, 13, or 26 mg / kg), Aprepitant alone (30, 50, 70 mg / kg), Ciclesonide alone (30 mg / kg loading and 2.54 mg / kg maintenance), Ciclesonide+Atosiban (11.8:1 FR^ 30 +2.54 mg / kg, 1.8 + 0.15 maintenance), Ciclesonide+Ivermectin (1.8:1 FR^ 30 + 16.7 mg / kg, 1.8 + 1mg / kg maintenance), Indomethacin+Nelfinavir Mesylate (1:3 FR^ mg / kg loading and maintenance doses for each drug will be determined) or Indomethacin+Ivermectin (1:7.7 FR^ mg / kg loading and maintenanceVU Ref.24027 / MBF Ref.093386-0031-WO01 doses for each drug will be determined). Mice (N ^ 5 per group) will receive twice daily (8 am and 8 pm) treatments of vehicle or test molecules on days 16–18. [000116] The onset of delivery will be monitored using infrared video surveillance and defined as delivery of the first pup. Preterm birth was defined as delivery occurring 24 hours prior to the average onset of term delivery of untreated mice. The number of pups and neonatal outcome (surviving pups born at term per total litter size including non-viable pups born, pups resorbed in utero, and pup weights at term delivery) will be recorded. [000117] Whether pups born at term from drug-treated dams are developmentally as mature as pups born to untreated dams will be determined by using reliable fetal growth markers relevant to human development, as previously noted (McCarthy et al., Biol Reprod.2018^ 99(5): 922-937). To do so, after delivery the pups from drug-treated and untreated (normal term) dams will be divided into two groups to assess skin or lung maturation. Similarly, skin barrier function in humans against infection and temperature / fluid loss occurs late in gestation (gestation day (gd) 17) and is fully formed at term. (i) Skin barrier function will be assessed by performing a whole-mount skin permeability assay as previously reported (Hardman et al., Development.1998^ 125(8): 1541-52). Briefly, neonates will be euthanatized and incubated in methanol for 5 minutes. After washing in PBS, neonates will be stained with 0.1% toluidine blue in PBS for 1 hour, washed, and then imaged. (ii) Morphometric assessment of lung airspaces will be performed in collaboration to reveal the degree of mesenchymal thinning that occurs after branching morphogenesis is complete (term pups), prior to alveolarization. To do so, neonatal lungs will be harvested, sectioned, H&E stained, and imaged (5–6 fields per lung). Computer-assisted morphometry will be performed using ImageJ to count the number of airspaces and measure distal airspace area (Benjamin et al., Clin Invest.2021^ 131(1)^ Benjamin et al., Am J Pathol.2016^ 186(7): 1786-800^ Plosa et al., Development.2014^ 141(24): 4751-62). Example 11 Clinical In Vivo Therapeutic Analysis of Preterm Birth [000118] The purpose of the project is to determine whether the preterm birth rate is reduced in moms taking one of the ten medications on our list that have been identified herein to have tocolytic potential for treatment of preterm labor. Additionally, the association between pregnancy exposure to any medication and reduced rates of preterm birth will be determined.VU Ref.24027 / MBF Ref.093386-0031-WO01 The clinical informatics core has extracted electronic health record data out of Epic’s Clarity database. [000119] Cohort: Adult women (>=18 years) who have delivered at VUMC. Exclusions: women who have had induced labor or have clinical or histological signs of intrauterine inflammation, bacterial vaginosis, chorioamnionitis, premature rupture of fetal membranes^ women with pre-eclampsia or multiple pregnancy^ scheduled or repeat cesarean deliveries prior to completed 37 weeks. Variables: gestational age at delivery^ age^ race^ BMI^ marital status^ education level^ smoking status^ infant birth weight^ tocolytic use during pregnancy (11 medications)^ tocolytic start / end dates^ and tocolytic doses. * * * [000120] The foregoing description of the specific aspects will so fully reveal the general nature of the invention that others can, by applying knowledge within the skill of the art, readily modify and / or adapt for various applications such specific aspects, without undue experimentation, without departing from the general concept of the present disclosure. Therefore, such adaptations and modifications are intended to be within the meaning and range of equivalents of the disclosed aspects, based on the teaching and guidance presented herein. It is to be understood that the phraseology or terminology herein is for the purpose of description and not of limitation, such that the terminology or phraseology of the present specification is to be interpreted by the skilled artisan in light of the teachings and guidance. [000121] The breadth and scope of the present disclosure should not be limited by any of the above-described exemplary aspects, but should be defined only in accordance with the following claims and their equivalents. [000122] All publications, patents, patent applications, and / or other documents cited in this application are incorporated by reference in their entirety for all purposes to the same extent as if each individual publication, patent, patent application, and / or other document were individually indicated to be incorporated by reference for all purposes. [000123] For reasons of completeness, various aspects of the invention are set out in the following numbered clauses:VU Ref.24027 / MBF Ref.093386-0031-WO01 [000124] Clause 1. A method for treating or preventing preterm labor or decreasing preterm birth in a pregnant subject in need thereof comprising administering to the subject a composition comprising a therapeutically effective amount of aprepitant or prodrug thereof. [000125] Clause 2. The method of clause 1, wherein the composition comprises a pharmaceutically acceptable carrier. [000126] Clause 3. The method of clause 1, wherein the prodrug is fosaprepitant. [000127] Clause 4. A method for treating or preventing preterm labor or decreasing preterm birth in a pregnant subject in need thereof comprising administering to the subject a composition comprising a therapeutically effective amount of ciclesonide. [000128] Clause 5. The method of clause 4, wherein the composition comprises a pharmaceutically acceptable carrier. [000129] Clause 6. The method of clause 4, further comprising administering atosiban to the subject. [000130] Clause 7. The method of clause 6, wherein the ciclesonide and the atosiban are administered to the subject at a ratio of about 11.8:1. [000131] Clause 8. The method of clause 6 or clause 7, wherein the ciclesonide and the atosiban are administered concomitantly. [000132] Clause 9. The method of clause 4, further comprising administering ivermectin to the subject. [000133] Clause 10. The method of clause 9, wherein the ciclesonide and the ivermectin are administered to the subject at a ratio of about 1.8:1. [000134] Clause 11. The method of clause 9 or clause 10, wherein the ciclesonide and the ivermectin are administered concomitantly. [000135] Clause 12. A method for treating or preventing preterm labor or decreasing preterm birth in a pregnant subject in need thereof comprising administering to the subject a composition comprising a therapeutically effective amount of ivermectin.VU Ref.24027 / MBF Ref.093386-0031-WO01 [000136] Clause 13. The method of clause 12, wherein the ivermectin is administered in the form of a composition, wherein the composition comprises a pharmaceutically acceptable carrier. [000137] Clause 14. The method of clause 12, further comprising administering indomethacin to the subject. [000138] Clause 15. The method of clause 14, wherein the ivermectin and the indomethacin are administered to the subject at a ratio of about 1:7.7. [000139] Clause 16. The method of clause 14 or clause 15, wherein the ivermectin and the indomethacin are administered concomitantly. [000140] Clause 17. A method for treating or preventing preterm labor or decreasing preterm birth in a pregnant subject in need thereof comprising administering to the subject a composition comprising a therapeutically effective amount of nelfinavir mesylate. [000141] Clause 18. The method of clause 17, wherein the composition comprises a pharmaceutically acceptable carrier. [000142] Clause 19. The method of clause 17, further comprising administering indomethacin to the subject. [000143] Clause 20. The method of clause 19, wherein the nelfinavir mesylate and the indomethacin are administered to the subject at a ratio of about 1:3. [000144] Clause 21. The method of clause 19 or clause 20, wherein the nelfinavir mesylate and the indomethacin are administered concomitantly. [000145] Clause 22. The method of any one of clauses 1-21, wherein the composition is administered orally, vaginally, rectally, subcutaneously, intraveneously, or by intramuscular injection. [000146] Clause 23. The method of any one of clauses 1-22, wherein the composition is administered once or more than once. [000147] Clause 24. The method of any one of clauses 1-23, wherein the composition is administered to the subject once or more than once per day.VU Ref.24027 / MBF Ref.093386-0031-WO01 [000148] Clause 25. The method of any one of clauses 1-24, wherein the method prevents the onset of preterm labor. [000149] Clause 26. The method of any one of clauses 1-25, wherein the composition is administered after the onset of preterm labor. [000150] Clause 27. The method of any one of clauses 1-26, wherein the composition is administered before the onset of preterm labor. [000151] Clause 28. The method of any one of clauses 1-27, wherein the subject exhibits one or more of the following before administration of the composition: recurrent uterine contractions^ cervical dilation^ cervical effacement^ and a cervical change. [000152] Clause 29. The method of any one of clauses 1-28, wherein the composition is administered with a secondary treatment for preterm labor. [000153] Clause 30. The method of clause 29, wherein the secondary treatment for preterm labor comprises nifedipine, indomethacin, magnesium sulfate, terbutaline, Ritodrine, Atosiban, Barusiban, nitroglycerin, Makena, hydroxyprogesterone caproate, progesterone supplementation, or cervical cerclage. [000154] Clause 31. The method of any one of clauses 1-30, wherein the composition is administered repeatedly beginning about the 18th to 22nd week of gestation until about the 37th week of gestation. [000155] Clause 32. The method of any one of clauses 1-30, wherein the composition is administered repeatedly for about 14 to 19 weeks. [000156] Clause 33. The method of any one of clauses 1-32, wherein the method further comprises monitoring or confirming a reduction in effacement, uterine contractions, cervical dilation, or a combination thereof after the administration of the composition. [000157] Clause 34. The method of any one of clauses 1-33, wherein the composition prolongs gestation. [000158] Clause 35. A method for treating or preventing preterm labor or decreasing preterm birth in a pregnant subject in need thereof comprising administering to the subject a compositionVU Ref.24027 / MBF Ref.093386-0031-WO01 comprising a therapeutically effective amount of aprepritant, fosaprepitant, ciclesonide, atosiban, ivermectin, indomethacin, nelfinavir mesylate, or a combination thereof.

Claims

VU Ref.24027 / MBF Ref.093386-0031-WO01 CLAIMS What is claimed is:

1. A method for treating or preventing preterm labor or decreasing preterm birth in a pregnant subject in need thereof comprising administering to the subject a composition comprising a therapeutically effective amount of aprepitant or prodrug thereof.

2. The method of claim 1, wherein the composition comprises a pharmaceutically acceptable carrier.

3. The method of claim 1, wherein the prodrug is fosaprepitant.

4. A method for treating or preventing preterm labor or decreasing preterm birth in a pregnant subject in need thereof comprising administering to the subject a composition comprising a therapeutically effective amount of ciclesonide.

5. The method of claim 4, wherein the composition comprises a pharmaceutically acceptable carrier.

6. The method of claim 4, further comprising administering atosiban to the subject.

7. The method of claim 6, wherein the ciclesonide and the atosiban are administered to the subject at a ratio of about 11.8:

1.

8. The method of claim 6, wherein the ciclesonide and the atosiban are administered concomitantly.

9. The method of claim 4, further comprising administering ivermectin to the subject.

10. The method of claim 9, wherein the ciclesonide and the ivermectin are administered to the subject at a ratio of about 1.8:

1.

11. The method of claim 9, wherein the ciclesonide and the ivermectin are administered concomitantly.VU Ref.24027 / MBF Ref.093386-0031-WO01 12. A method for treating or preventing preterm labor or decreasing preterm birth in a pregnant subject in need thereof comprising administering to the subject a composition comprising a therapeutically effective amount of ivermectin.

13. The method of claim 12, wherein the ivermectin is administered in the form of a composition, wherein the composition comprises a pharmaceutically acceptable carrier.

14. The method of claim 12, further comprising administering indomethacin to the subject.

15. The method of claim 14, wherein the ivermectin and the indomethacin are administered to the subject at a ratio of about 1:7.

7.

16. The method of claim 14, wherein the ivermectin and the indomethacin are administered concomitantly.

17. A method for treating or preventing preterm labor or decreasing preterm birth in a pregnant subject in need thereof comprising administering to the subject a composition comprising a therapeutically effective amount of nelfinavir mesylate.

18. The method of claim 17, wherein the composition comprises a pharmaceutically acceptable carrier.

19. The method of claim 17, further comprising administering indomethacin to the subject.

20. The method of claim 19, wherein the nelfinavir mesylate and the indomethacin are administered to the subject at a ratio of about 1:

3.

21. The method of claim 19, wherein the nelfinavir mesylate and the indomethacin are administered concomitantly.

22. The method of claim 1, wherein the composition is administered orally, vaginally, rectally, subcutaneously, intraveneously, or by intramuscular injection.VU Ref.24027 / MBF Ref.093386-0031-WO01 23. The method of claim 1, wherein the composition is administered once or more than once.

24. The method of claim 1, wherein the composition is administered to the subject once or more than once per day.

25. The method of claim 1, wherein the method prevents the onset of preterm labor.

26. The method of claim 1, wherein the composition is administered after the onset of preterm labor.

27. The method of claim 1, wherein the composition is administered before the onset of preterm labor.

28. The method of claim 1, wherein the subject exhibits one or more of the following before administration of the composition: recurrent uterine contractions^ cervical dilation^ cervical effacement^ and a cervical change.

29. The method of claim 1, wherein the composition is administered with a secondary treatment for preterm labor.

30. The method of claim 29, wherein the secondary treatment for preterm labor comprises nifedipine, indomethacin, magnesium sulfate, terbutaline, Ritodrine, Atosiban, Barusiban, nitroglycerin, Makena, hydroxyprogesterone caproate, progesterone supplementation, or cervical cerclage.

31. The method of claim 1, wherein the composition is administered repeatedly beginning about the 18th to 22nd week of gestation until about the 37th week of gestation.

32. The method of claim 1, wherein the composition is administered repeatedly for about 14 to 19 weeks.VU Ref.24027 / MBF Ref.093386-0031-WO01 33. The method of claim 1, wherein the method further comprises monitoring or confirming a reduction in effacement, uterine contractions, cervical dilation, or a combination thereof after the administration of the composition.

34. The method of claim 1, wherein the composition prolongs gestation.

35. A method for treating or preventing preterm labor or decreasing preterm birth in a pregnant subject in need thereof comprising administering to the subject a composition comprising a therapeutically effective amount of aprepritant, fosaprepitant, ciclesonide, atosiban, ivermectin, indomethacin, nelfinavir mesylate, or a combination thereof.

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