NT-NOA - Novel Treatment of Some Men With Non-Obstructive Azoospermia

This clinical interventional study aims to assess whether treatment with denosumab in combination with letrozole can improve sperm production in infertile men with non-obstructive azoospermia selected by serum anti-mullerian hormone (AMH) as a positive predictive biomarker.

Infertility is a health issue worldwide with more than 15% of couples experiencing infertility. Male factors contribute in 50% of all cases and is the sole cause in 30% of all infertile couples . Treatment options for the male infertility are limited and only specific conditions like varicocele and hypogonadotropic hypogonadism can be effectively treated with surgery and hormonal stimulation, respectively. However, for the remaining 90% of infertile men, no treatment options exist. Instead, their female partners must undergo fertility treatment.

Men with the most pronounced impairment of semen quality, men with azoospermia, have no sperm in the ejaculate and are classically divided into obstructive azoospermia (OA) and non-obstructive azoospermia (NOA), with the ladder being the most common. These men needs to have surgery performed for sperm retrieval with either testicular sperm aspiration (TESA), testicular sperm extraction (TESE) or micro testicular sperm extraction (microTESE) in order to have a chance of becoming biological fathers. The chance of successful sperm retrieval in men with OA is close to 100%, whereas men with NOA have a significantly lower chance with only around 10-40% being successful. This is due to the vast difference in etiology between men with OA and NOA. Even with a successful surgical intervention only half of the cases will result in a live birth, leaving the majority of men with NOA no chance of becoming biological fathers. Therefore, novel treatment options for even a minor sub-group of these men are warranted.

RANKL is a ligand for the receptor activator of nuclear factor kappa beta (RANK), and their pathway plays a prominent role in the regulation of bone metabolism. The binding of RANKL to RANK on osteoclast precursors induces osteoclast maturation and activation, thereby stimulating bone resorption, and regulates cell cycle i.e., proliferation, differentiation, and apoptosis. Osteoprotegerin (OPG) is a secreted decoy receptor that controls RANKL-RANK interaction by binding RANKL and inhibits activation of RANK and preventing osteoclast differentiation and activation.

Denosumab, a drug used in millions of patients worldwide under trade names Prolia® and Xgeva®, inhibits the RANKL pathway and is used to treat osteoporosis and bone metastases. The drug's mechanism of action inhibits RANKL and thus inhibits bone resorption through reduced osteoclast activation. This reduces the loss of bone mineral density (BMD), which reduces the risk of bone loss and thereby the risk of fracture and osteoporosis. Denosumab has been shown in several clinical studies to be a safe and effective drug in both women and men and has been in clinical use in both sexes for many years. As Denosumab has a teratogenic effect, pharmacokinetic studies in both monkeys and healthy men were performed before approval of the drug as a treatment for osteoporosis in men. These studies showed that Denosumab concentration in semen does not pose a risk to the fetus during sexual intercourse with the pregnant woman and therefore is safe to use for the suggested infertility indication as there is no risk of fetal transmission.

The investigators have also shown that RANKL also influences germ cell apoptosis, differentiation, and proliferation in mice and humans. This indicates that the RANKL system could be a potential direct regulator of spermatogenesis, and thus highlighting that the drug Denosumab, a RANKL inhibitor, commonly used to treat osteoporosis, may be used for medical treatment of men with decreased sperm production. The effects of denosumab in vitro were supported by a pilot intervention study in 12 infertile men, which showed that denosumab either had a highly beneficial or detrimental effect on sperm production. This implies that only a sub-group of infertile men should be offered this treatment and these men should be identified by, preferably, a simple and accessible biomarker.

Therefore, the investigators are currently conducting two randomized clinical trials "First In Treating Infertility (FITMI)" and "Novel Approach for Oligospermia (NAPO) to explore whether denosumab can improve semen quality in infertile men selected by serum AMH, but already have sperm concentration between 2 mio/mL - 20 mio/mL (FITMI) and 0.01 mio/mL - 2 mio/mL (NAPO). These are both important studies, but still leaves out men with no sperm in the ejaculate, azoospermic patients. Therefore, there is a need for a trial adressing these patients for a potential novel treatment option.

NT-NOA is a single-center, sponsor-investigator-initiated, interventional trial, Following successful completion of screening procedures, subjects will all receive denosumab 60 mg s.c. and Letrozole oral tablet 2.5 mg once daily. Participants will all initiate letrozole treatment 14 days prior to the denosumab injection and will continue until the TESE procedure. The study will be carried out at the Division of Translational Endocrinology, Copenhagen University Hospital, Herlev, Copenhagen, Denmark.

The investigators believe it is of clinical relevance to treat 10 patients with denosumab in combination with letrozole for 1 of them to achieve sperm production, however, it is difficult to estimate. Ideally 50% of the treated participants will have sperm suitable for ART post intervention. By including 16 patients, there will be sufficient power to determine if combination therapy with denosumab and letrozole induces a clinically relevant effect on spermatogenesis.

Eligibility Criteria:
Inclusion Criteria:

* Infertile men ≥ 18 years and \< 60 years of age with azoospermia with normal semen volume (\> 0.9 mL) verified by minimum 2 semen samples.
* Testis size (Prader Orchidometer) ≤ 15 mL on both sides.
* No identifiable cause of obstruction.
* Serum AMH levels ≥5 pmol/L.

Exclusion Criteria:

* Chronic diseases, defined as diagnosis where signs, symptoms, and treatment imply an expected long duration and lack of a cure, such as diabetes mellitus, metabolism disorders, osteoporosis, colitis, etc.
* Chromosome anomalies e.g. Klinefelter Syndrome, XX male
* AZFa/b/c microdeletions besides b2/b3 og gr/gr
* Detected CFTR mutation
* Men with current or previous malignancies, or at potential risk of testicular cancer after baseline examination and ultrasound will be excluded
* Men with hypocalcemia at baseline, defined as albumin corrected calcium \< 2.17 mmol/L or total calcium \< 2.14 mmol/L
* Serum vitamin D (25OHD) levels \< 25 nmol/L
* estimated Glomerular Filtration Rate (eGFR) \< 60 mL/min/1,73 m2
* Insufficient dental status
* Hypersensitivity to latex, Denosumab, or to any of the excipients (acetic acid, sodium hydroxide, Sorbitol (E420), Polysorbate 20) will be excluded.
* Serum Inhibin B \< 5 pg/mL
* Androgen replacement therapy
* Hypogonadotropic hypogonadism
* BMI ≥ 35 kg/m2