The Effect of the Probiotic Strains on Bone Mineral Density in Postmenopausal Women

The main purpose of the study is to assess the effect of oral probiotic supplementation on bone mineral density in postmenopausal women. A total of 170 female subjects who meet all the inclusion criteria and none of the exclusion criteria are to be included and randomized to one of two arms (the probiotic arm or the placebo arm) at a ratio of 1:1.

The subjects who provide their written informed consent will receive oral probiotic formulation containing a mixture of two strains Lactobacillus plantarum and Lactobacillus paracasei or placebo. The probiotic formulation and placebo will be provided by Nordic Biotic Ltd. (the company will also ensure study drug blinding). Study subjects will take the provided probiotic formulation/placebo orally once daily for 12 months.

The study timeline spans 54 weeks and involves 13 visits, including:

• Screening visit - subjects eligibility to participate in the study will be assessed based on the inclusion/exclusion criteria - the screening period will last up to 14 before the study drug is administered;
• Randomization visit (visit 0) - subjects will be randomly allocated to study groups and will receive a supply of the study probiotic or placebo;
• 6 office visits (at months 2, 4, 6, 8, 10, and 12 of visit 0);
• 5 remote visits (at months 1, 3, 7, 9, and 11 of visit 0).

All subjects included in the study will undergo:

• History-taking and physical examination at each office visit,
• Nutritional status assessment (measurements of body weight and height, with a BMI calculation) at the screening visit and at months 6 and 12 of the study intervention,
• Two densitometry scans (DEXA) of the lumbar segment of the spine (L1-L4) at the screening visit and at month 12 of visit 0,
• Treatment satisfaction assessment (treatment satisfaction questionnaire) at months 2, 6, 10, and 12 of the study intervention.

At each visit, whether conducted at the doctor's office or remotely, subjects will be asked about their use of the study drug and any gastrointestinal symptoms (number of bowel movements, stool consistency, bloating, abdominal pain), and the use of any other medications (including antibiotics). Each time, the subjects will be also asked about any side effects. Stool consistency will be assessed with the Bristol Stool Form Scale, and abdominal pain, bloating, and nausea will be assessed with a 5-grade Likert scale.

Blood samples will be collected from all subjects during the screening period, after 6 months of intervention and at the completion of intervention (i.e. at month 12).

The following blood tests will be conducted:

• Complete blood count, erythrocyte sedimentation rate, and C-reactive protein levels (at screening and at months 6 and 12 of the study intervention);
• Serum calcium, phosphorus, and alkaline phosphatase levels (at screening and at months 6 and 12 of the study intervention);
• Serum vitamin D and parathormone levels (at screening and at month 12 of intervention).

Inclusion criteria

• Postmenopausal women (between 2 and 5 years after their last menstrual period)
• A dual-energy X-ray absorptiometry (DEXA) scan-based T-score of the lumbar spine (L1-L4) of less than -1.49 (i.e. a normal value according to the World Health Organization)
• Body mass index (BMI) between 18 and 30
• Vitamin 25 (OH)D levels between 31-50 ng/mL

Exclusion criteria

• The last menstrual period less than 2 or more than 5 years before study recruitment
• BMI under 18 or above 30
• Oral or transdermal hormone therapy with estrogens and/or progestogens within the last 6 months
• Corticosteroid or thyroid hormone therapy within the last 6 months
• History of osteopenia or osteoporosis
• Neoplastic disease treated with cytostatic agents within the last 12 months
• Autoimmune disorders, including rheumatoid arthritis, systemic lupus erythematosus, ankylosing spondylitis; malabsorption syndromes, including celiac disease; chronic kidney disease; chronic liver failure; endocrine disorders, including Hashimoto thyroiditis, Graves' disease, Cushing disease or syndrome, hyperparathyroidism, hypogonadism, hyperprolactinemia, acromegaly, diabetes mellitus; and other chronic conditions affecting bone metabolism
• Antibiotic therapy within the last 2 months prior to study recruitment
• Probiotic use within the last 2 months prior to study recruitment
• Calcium and vitamin D supplementation within the 2 months prior to study recruitment
• Treatment with antidepressant or antipsychotic agents involving a change of the agents or dosage within the last 3 months
• Substance use disorder (alcohol, drugs, nicotine)
• History of organ transplantation
• Exposure to the SARS-Cov-2 virus within 14 days prior to study recruitment (based on patient declaration)
• An acute COVID-19 infection within 14 days prior to study recruitment (based on patient declaration)
• A surgical procedure scheduled to take place during this study
• Participation in another study within the last 6 months
• Inability to give an informed consent in writing
• Any disease or circumstances that, according to the Investigator or Sponsor, might prevent the subject from completing the study or following study procedures and requirements