Sex Hormones and Atherosclerosis Prevention in Perimenopausal Women
Condition: Menopause; Aging
Study type: Interventional
Phase: Early Phase 1

Summary: 
      As women get older and go through menopause, levels of the female reproductive hormone
      estradiol decrease to low levels. Also with aging, the functioning of the arteries declines.
      Over time this vascular dysfunction can lead to health problems such as high blood pressure
      and heart disease. This study is being done to help determine what causes arteries to become
      unhealthy in postmenopausal women, who have low levels of the female reproductive hormone
      estradiol. In this study we will test whether low levels of tetrahydrobiopterin (BH4), a
      natural substance in the body that can cause the arteries to expand, explains why arteries
      become unhealthy in women with low levels of estradiol. To answer this question, we will
      study how vascular function changes with a medication that causes a short-term increase in
      BH4 levels when estradiol is lowered with a medication, compared to when estradiol is normal.
      We will also determine whether the administration of the antioxidant vitamin C, along with
      the medication to increase BH4 levels, will normalize vascular health in perimenopausal and
      postmenopausal women, and in women who have their estradiol levels lowered.
    

Detailed description: 
      Endothelial dysfunction, featuring reduced vascular endothelial-dependent dilation (EDD), is
      a significant predictor of cardiovascular (CV) events. Reduced EDD is characterized by the
      loss of nitric oxide (NO) production. EDD declines with aging in women at a slower rate
      compared to men but rapidly declines after menopause and quickly catches up to men,
      suggesting a beneficial effect of estrogen (E2). Our research has investigated the mechanisms
      underlying endothelial protection of E2 including how hormonal fluctuations during the
      peri-to early postmenopausal years contribute to endothelial dysfunction, and how these
      hormonal changes might interact with aging. Using a well described experimental model
      (systemic infusion of ascorbic acid; AA) to acutely reduce reactive oxygen species (ROS), our
      preliminary findings implicate oxidative stress as an underlying mechanism explaining the
      impaired EDD during the menopausal transition. Specifically we found improved EDD in response
      to AA in late peri- and postmenopausal women but not in pre- or early perimenopausal women.
      These data indicate that E2 may protect the vasculature from free radical injury during pre-
      and early perimenopausal years. In support of this concept, we demonstrated that AA can
      reverse the decrease in EDD that occurs in the setting of short-term E2 deficiency (3-days of
      gonadotropin releasing hormone antagonist; GnRHant) in pre-and early perimenopausal women.

      Tetrahydrobiopterin (BH4) is a critical co-factor in the production of NO by endothelial
      nitric oxide synthase (eNOS), and in the face of deficient levels, uncoupling of eNOS results
      in the production of ROS instead of NO. Whether the increased ROS and impaired EDD that
      occurs during the peri- and early postmenopausal periods is associated with reduced vascular
      BH4 is at present unknown. However, our finding that AA could not improve the reduced EDD
      found in early perimenopausal women or restore EDD in peri-and postmenopausal women to
      premenopausal levels, indicates that other sources of ROS or causes of low NO are involved in
      the impaired EDD across the stages of the menopause transition and aging in women. In vitro
      studies demonstrate that the co-administration of BH4 and AA completely prevents the eNOS
      uncoupling and decreased NO production in endothelial cells treated with a peroxynitrite
      (ONOO-) donor, compared to administering either one alone. The global aim of this renewal is
      to follow-up on our previous findings and to determine the role of BH4 as a mediator of
      endothelial dysfunction [across the stages of the menopausal transition and aging in women.
      This global aim will be addressed by 3 specific aims (SA). Using a cross-sectional design,
      SA1 will measure EDD in healthy pre-, peri- and postmenopausal women: 1) under basal
      conditions; and 2) following an acute increase in BH4. SA2 will expand on the cross-sectional
      comparisons via short-term (10 days) ovarian suppression (GnRhant) in pre-and early
      perimenopausal women to isolate the age-independent effects of E2. SA3 will examine whether
      co-administration of BH4 and AA fully restore youthful EDD in peri-and postmenopausal women.

      SA1: To determine the possible mechanistic involvement of BH4 deficiency in the decline in
      endothelial function during the peri-and early postmenopausal period and its potential
      relation to oxidative stress. This will be assessed by comparing brachial artery EDD and
      plasma and endothelial cell markers of oxidative stress and NO (see methods for details) in
      healthy pre-, peri-and postmenopausal women: 1) under basal conditions; and 2) in response to
      acute oral BH4 (vs PL) supplementation.

      Hypothesis1 (H1): a) EDD will increase in response to BH4 in early peri, late peri and
      postmenopausal women but not premenopausal women. b) The increase in EDD in response to BH4
      will be associated with a decrease in plasma markers of oxidative stress. c) The reduced EDD
      in peri- and postmenopausal women will be associated with lower endothelial cell markers of
      NO and higher oxidative Stress.

      SA2: To determine whether the decrease in EDD in response to acute ovarian suppression in
      pre- and early perimenopausal women is related to reduced BH4 bioavailability and elevated
      markers of oxidative stress. This will be assessed by comparing EDD and plasma markers of
      oxidative stress after acute oral supplementation of BH4 (or PL) following either acute
      ovarian suppression with (GnRHant+E2) or without (GnRHant+PL) E2 add-back. Additionally,
      endothelial cell protein expression of markers of BH4 biosynthesis and oxidative stress will
      be measured before and after the suppression/add-back paradigm.

      H2: a) The expected decrease in EDD and increase in plasma oxidative stress markers following
      acute ovarian suppression will be mitigated in the groups treated with oral BH4
      (GnRHant+PL+BH4); there will be little or no effect of the addition of BH4 to the E2 add-back
      group. b) Ovarian suppression alone will reduce endothelial protein markers of BH4
      biosynthesis and will increase markers of oxidative stress. These changes will not be
      observed following ovarian suppression with E2 add-back and oral BH4. These sets of
      experiments will provide evidence that the oxidative stress-related decrease in EDD in
      response to E2 suppression is in part, related to decreased BH4 bioavailability.

      SA3: To determine whether the co-administration of BH4 with AA will completely restore EDD in
      peri-and postmenopausal women, and in early perimenopausal women following ovarian
      suppression.

      H3: The co-administration of BH4 with AA will restore EDD to premenopausal levels a) in
      peri-and postmenopausal women, and b) in early perimenopausal following acute ovarian
      suppression.
    

Intervention name(s): KUVAN; Placebo; Cetrotide; Climara; Placebo transdermal patch
Intervention type(s): Drug; Drug; Drug; Drug; Drug
Intervention description(s): KUVAN will be given orally at 10mg/kg dose per body weight. KUVAN will be dissolved in 4-8oz water and consumed within 15 minutes.; Placebo pills designed to match the dissolvable KUVAN pills; Cetrotide (0.25mg/d) will be taken daily for 10 days via abdominal subcutaneous injection.; 0.075mg/d transdermal patch will be placed on skin.; Inactive transdermal patch
Other intervention name(s): Tetrahydrobiopterin or BH4; Control; Cetrorelix acetate; Estradiol transdermal patch; Control
Intervention MeSH terms: Cetrorelix; Estradiol
Allocation: Randomized
Intervention model: Parallel Assignment
Masking: Quadruple (Participant, Care Provider, Investigator, Outcomes Assessor)
Primary purpose: Basic Science
Keywords: endothelial function; women; estrogen deficiency; sex hormones; adiposity; oxidative stress; antioxidants