Fertility, Contraception
Conditions
Keywords
Contraception, Mucus, Cervix, progestin, hormones
Brief summary
The purpose of this study is to help us determine how hormones impact the production of cervical mucus which is a fluid secreted by the opening of the uterus, also called the cervix. During a normal menstrual cycle, cervical mucus changes in response to two hormones that the ovaries make: estrogen and progesterone. Many of the birth control methods women use thicken mucus and make it less likely for sperm to enter the uterus. The goal of this study is to better understand the effects estrogen and progesterone have on the cervical mucus in the body. To do this, the investigators will be using a drug that suppresses natural hormone production for 3 months. Estrogen and progesterone will be artificially replaced to better understand their effects. The investigators will also be studying the effect of a common birth control pill that may work by thickening cervical mucus. Last, the investigators will study some of the cells in the cervix that produce mucus.
Detailed description
This proposed study is to evaluate the temporal changes in human cervical mucus in response to progesterone and progestins as an initial step toward clarifying these relationships. Our hypothesis is that direct effects of progesterone on the endocervix, independent of estrogen withdrawal, cause contraceptive changes to cervical mucus. To study this, a randomized, prospective, crossover study will be conducted by examining cervical mucus changes in a small cohort of women in whom investigators will suppress circulating hormonal levels by administering a GnRH antagonist. Investigators will then artificially replace Estrogen and Progesterone in order to differentiate their effects on clinical and laboratory measurements of mucus quality. The investigators will be looking closely at the immediate changes in mucus when .35 mg of norethindrone, a marketed drug is administered in this experimental setting. Cervical cell samples will also be collected at various time points and perform RT-PCR to determine whether genes for membrane bound progesterone receptor are expressed and regulated by Estrogen and Progesterone.
Interventions
DRUGLeuprolide acetate
Following this spontaneous cycle, participants received a single intramuscular injection of leuprolide acetate (11.25mg, LupronVR; Abbvie, Chicago, IL), a dose documented to suppress ovarian function for at least 3 months. Ovarian suppression is confirmed with serum E2 less than or equal to 35 pg/ml 21-28 day post-injection.
DRUGEstrogen and Progesterone Replacement
Replace hormones exogenously to create an artificial cycle
DRUGProgestin
Synthetic progestin to assess effects on cervical mucus.
Sponsors
Society of Family Planning
Oregon Health and Science University
Study design
Allocation
RANDOMIZED
Intervention model
CROSSOVER
Primary purpose
BASIC_SCIENCE
Masking
SINGLE (Investigator)
Eligibility
Sex/Gender
FEMALE
Age
21 Years to 40 Years
Healthy volunteers
Yes
Inclusion criteria
* Normal menstrual cycles of 25-35 days in length for at least previous 3 cycles * 21-40 years of age * BMI \>18, \<30 * Serum P4 ≥ 3 ng/ml on single sample collected between days 18-25 of self-reported menstrual cycle * Flexible schedule allowing morning blood draws on less than 48 hour notice * In good general health * Commit to remain on stable diet during study period (no changes to normal dietary habits) * Commit to using non-hormonal contraceptive methods during study period except those prescribed in the experimental protocol * No objections to taking study drugs * No objections to refraining from intercourse the night before any sampling and willing to using condoms during vaginal intercourse.
Exclusion criteria
* Oral contraceptive use or other hormone supplement within the preceding 2 months * Women with current cervical infection * Evidence of abnormal cervical cytology * Use of Paragard IUD for contraception * Long-acting hormonal contraceptive use in the past 12 months (e.g., Depo-Provera®) * Contraindications to study drugs * Current or past pregnancy within the previous 6 months or currently trying to conceive * Desiring to conceive in the next 8 months * Breastfeeding in the past 2 months * Diagnosed Diabetes or Metabolic Syndrome * Current or previous use of cholesterol lowering drugs within the preceding 12 months * Diagnosed Polycystic Ovary Syndrome * History of, or self-reported, substance abuse * Smoker * Previous infertility treatment excluding male factor issues * Use of an investigational drug within the past 2 months * History of excisional or ablative treatment procedure on cervix (ie. LEEP, Cryotherapy, Cold Knife Cone)
Design outcomes
Primary
| Measure | Time frame | Description |
|---|---|---|
| Median Cervical Mucus Score - Baseline | Baseline | Measurement of median cervical mucus scores at baseline. The current clinical standard for appraising cervical mucus is the cervical mucus score (ie. Insler score) that examines mucus based on 5 metrics including volume, spinnbarkeit (stretch), ferning, viscosity and cellularity on a 15-point scale. Per WHO guidelines, scores above 10 are considered mucus favoring penetration and scores below 10 are considered to be unfavorable to penetration. |
| Sperm Penetration Scores | Approximately one year | Measuring sperm penetration scores in different hormonal conditions |
| Median Cervical Mucus Score - 2 Hour | 2 hours | Measurement of median cervical mucus scores 2 hours following norethindrone administration. The current clinical standard for appraising cervical mucus is the cervical mucus score (ie. Insler score) that examines mucus based on 5 metrics including volume, spinnbarkeit (stretch), ferning, viscosity and cellularity on a 15-point scale. Per WHO guidelines, scores above 10 are considered mucus favoring penetration and scores below 10 are considered to be unfavorable to penetration. |
| Median Cervical Mucus Score - 6 Hour | 6 hours | Measurement of median cervical mucus scores 6 hours following norethindrone administration. The current clinical standard for appraising cervical mucus is the cervical mucus score (ie. Insler score) that examines mucus based on 5 metrics including volume, spinnbarkeit (stretch), ferning, viscosity and cellularity on a 15-point scale. Per WHO guidelines, scores above 10 are considered mucus favoring penetration and scores below 10 are considered to be unfavorable to penetration. |
| Median Cervical Mucus Score - 24 Hour | 24 hours | Measurement of median cervical mucus scores 24 hours following norethindrone administration. The current clinical standard for appraising cervical mucus is the cervical mucus score (ie. Insler score) that examines mucus based on 5 metrics including volume, spinnbarkeit (stretch), ferning, viscosity and cellularity on a 15-point scale. Per WHO guidelines, scores above 10 are considered mucus favoring penetration and scores below 10 are considered to be unfavorable to penetration. |
Secondary
| Measure | Time frame | Description |
|---|---|---|
| Change in PGRMC1 During Menstrual Cycle | 1 month | Measuring the mean transcript change of membrane bound progestin receptors (PGRMC1) from follicular phase to ovulation to luteal phase of spontaneous menstrual cycle. Total RNA (ribonucleic acid) was isolated from endocervical cell samples and analyzed for expression of PGRMC1 using real-time PCR (polymerase chain reaction) relative to levels of ribosomal (S10) RNA. An endocervical brush will be inserted into the os and then immediately rinsed into a special RNA preserving reagent. After total RNA is isolated and purified, it will be reverse transcribed into cDNA using primers. Ratio of PGRMC1 to 18s RNA Gene expression of membrane bound progesterone receptors in endocervical cells |
Countries
United States
Participant flow
Pre-assignment details
In order to qualify for the intervention phase, subjects needed to demonstrate an ovulatory menstrual cycle during the Screening Period.
Participants by arm
| Arm | Count |
|---|---|
| Norethindrone, Then Estradiol Withdrawal This arm will receive Norethindrone (NET) first then experience estradiol withdrawal (E2WD).
On the day prior to each admission, we instructed subjects to apply three fresh estradiol (E2) patches (0.3mg/day) to achieve mid-cycle level E2 effects.
On the day of the first and third admission, subjects were randomized to the experimental condition, either: (1) oral progestin administration (e.g., a single dose of 0.35mg norethindrone) with continuation of the E2 patches; or (2) estradiol withdrawal (e.g., removal of the patches). Subject then received the converse intervention for admissions two and four.
Subjects resumed treatment with the 0.1mg E2 patch until their next scheduled inpatient assessment. We required a minimum of five days at this level before repeating the dose escalation and the next evaluation. | 2 |
| Estradiol Withdrawal, Then Norethindrone This arm will experience estradiol withdrawal (E2WD) first and then receive Norethindrone (NET).
On the day prior to each admission, we instructed subjects to apply three fresh estradiol (E2) patches (0.3mg/day) to achieve mid-cycle level E2 effects.
On the day of the first and third admission, subjects were randomized to the experimental condition, either: (1) oral progestin administration (e.g., a single dose of 0.35mg norethindrone) with continuation of the E2 patches; or (2) estradiol withdrawal (e.g., removal of the patches). Subject then received the converse intervention for admissions two and four.
Subjects resumed treatment with the 0.1mg E2 patch until their next scheduled inpatient assessment. We required a minimum of five days at this level before repeating the dose escalation and the next evaluation. | 2 |
| Total | 4 |
Withdrawals & dropouts
| Period | Reason | FG000 | FG001 | FG002 |
|---|---|---|---|---|
| Screening Period | Hormones not suppressed post Lupron | 1 | 0 | 0 |
Baseline characteristics
| Characteristic | Norethindrone, Then Estradiol Withdrawal | Estradiol Withdrawal, Then Norethindrone | Total |
|---|---|---|---|
| Age, Categorical <=18 years | 0 Participants | 0 Participants | 0 Participants |
| Age, Categorical >=65 years | 0 Participants | 0 Participants | 0 Participants |
| Age, Categorical Between 18 and 65 years | 2 Participants | 2 Participants | 4 Participants |
| Race (NIH/OMB) American Indian or Alaska Native | 0 Participants | 0 Participants | 0 Participants |
| Race (NIH/OMB) Asian | 0 Participants | 0 Participants | 0 Participants |
| Race (NIH/OMB) Black or African American | 0 Participants | 0 Participants | 0 Participants |
| Race (NIH/OMB) More than one race | 0 Participants | 0 Participants | 0 Participants |
| Race (NIH/OMB) Native Hawaiian or Other Pacific Islander | 0 Participants | 0 Participants | 0 Participants |
| Race (NIH/OMB) Unknown or Not Reported | 0 Participants | 0 Participants | 0 Participants |
| Race (NIH/OMB) White | 2 Participants | 2 Participants | 4 Participants |
| Region of Enrollment United States | 2 participants | 2 participants | 4 participants |
| Sex: Female, Male Female | 2 Participants | 2 Participants | 4 Participants |
| Sex: Female, Male Male | 0 Participants | 0 Participants | 0 Participants |
Adverse events
| Event type | EG000 affected / at risk | EG001 affected / at risk | EG002 affected / at risk |
|---|---|---|---|
| deaths Total, all-cause mortality | 0 / 5 | 0 / 4 | 0 / 4 |
| other Total, other adverse events | 0 / 5 | 0 / 4 | 0 / 4 |
| serious Total, serious adverse events | 0 / 5 | 0 / 4 | 0 / 4 |
Outcome results
Primary
Median Cervical Mucus Score - 24 Hour
Measurement of median cervical mucus scores 24 hours following norethindrone administration. The current clinical standard for appraising cervical mucus is the cervical mucus score (ie. Insler score) that examines mucus based on 5 metrics including volume, spinnbarkeit (stretch), ferning, viscosity and cellularity on a 15-point scale. Per WHO guidelines, scores above 10 are considered mucus favoring penetration and scores below 10 are considered to be unfavorable to penetration.
Time frame: 24 hours
Population: Data from one (1) subject were excluded from analyses because estradiol (E2) levels were consistent with patch noncompliance
| Arm | Measure | Value (MEDIAN) |
|---|---|---|
| Norethindrone | Median Cervical Mucus Score - 24 Hour | 8 score on a scale |
| Estradiol | Median Cervical Mucus Score - 24 Hour | 10.5 score on a scale |
Primary
Median Cervical Mucus Score - 2 Hour
Measurement of median cervical mucus scores 2 hours following norethindrone administration. The current clinical standard for appraising cervical mucus is the cervical mucus score (ie. Insler score) that examines mucus based on 5 metrics including volume, spinnbarkeit (stretch), ferning, viscosity and cellularity on a 15-point scale. Per WHO guidelines, scores above 10 are considered mucus favoring penetration and scores below 10 are considered to be unfavorable to penetration.
Time frame: 2 hours
Population: Data from one (1) subject were excluded from analyses because estradiol (E2) levels were consistent with patch noncompliance
| Arm | Measure | Value (MEDIAN) |
|---|---|---|
| Norethindrone | Median Cervical Mucus Score - 2 Hour | 8.5 score on a scale |
| Estradiol | Median Cervical Mucus Score - 2 Hour | 10.5 score on a scale |
Primary
Median Cervical Mucus Score - 6 Hour
Measurement of median cervical mucus scores 6 hours following norethindrone administration. The current clinical standard for appraising cervical mucus is the cervical mucus score (ie. Insler score) that examines mucus based on 5 metrics including volume, spinnbarkeit (stretch), ferning, viscosity and cellularity on a 15-point scale. Per WHO guidelines, scores above 10 are considered mucus favoring penetration and scores below 10 are considered to be unfavorable to penetration.
Time frame: 6 hours
Population: Data from one (1) subject were excluded from analyses because estradiol (E2) levels were consistent with patch noncompliance
| Arm | Measure | Value (MEDIAN) |
|---|---|---|
| Norethindrone | Median Cervical Mucus Score - 6 Hour | 6.5 score on a scale |
| Estradiol | Median Cervical Mucus Score - 6 Hour | 8.5 score on a scale |
Primary
Median Cervical Mucus Score - Baseline
Measurement of median cervical mucus scores at baseline. The current clinical standard for appraising cervical mucus is the cervical mucus score (ie. Insler score) that examines mucus based on 5 metrics including volume, spinnbarkeit (stretch), ferning, viscosity and cellularity on a 15-point scale. Per WHO guidelines, scores above 10 are considered mucus favoring penetration and scores below 10 are considered to be unfavorable to penetration.
Time frame: Baseline
Population: Data from one (1) subject were excluded from analyses because estradiol (E2) levels were consistent with patch noncompliance
| Arm | Measure | Value (MEDIAN) |
|---|---|---|
| Norethindrone | Median Cervical Mucus Score - Baseline | 11 score on a scale |
| Estradiol | Median Cervical Mucus Score - Baseline | 13 score on a scale |
Primary
Sperm Penetration Scores
Measuring sperm penetration scores in different hormonal conditions
Time frame: Approximately one year
Population: Outcome not analyzed.
Secondary
Change in PGRMC1 During Menstrual Cycle
Measuring the mean transcript change of membrane bound progestin receptors (PGRMC1) from follicular phase to ovulation to luteal phase of spontaneous menstrual cycle. Total RNA (ribonucleic acid) was isolated from endocervical cell samples and analyzed for expression of PGRMC1 using real-time PCR (polymerase chain reaction) relative to levels of ribosomal (S10) RNA. An endocervical brush will be inserted into the os and then immediately rinsed into a special RNA preserving reagent. After total RNA is isolated and purified, it will be reverse transcribed into cDNA using primers. Ratio of PGRMC1 to 18s RNA Gene expression of membrane bound progesterone receptors in endocervical cells
Time frame: 1 month
Population: PGRMC1 data was analyzed for the 4 subjects who demonstrate ovarian suppression following leuprolide injection and continued to intervention phase of the study. Outcome was only evaluated during spontaneous menstrual cycle (1 month).~One participant corresponds to one cycle.
| Arm | Measure | Group | Value (MEAN) | Dispersion |
|---|---|---|---|---|
| Norethindrone | Change in PGRMC1 During Menstrual Cycle | Follicular Phase | 1.2 ratio | Standard Deviation 0.38 |
| Norethindrone | Change in PGRMC1 During Menstrual Cycle | Ovulation Phase | 4.0 ratio | Standard Deviation 2.4 |
| Norethindrone | Change in PGRMC1 During Menstrual Cycle | Luteal Phase | 1.02 ratio | Standard Deviation 0.73 |