Surgical Sperm Retrieval in Non-obstructive Azoospermic Men: mTESE vs. TESA
Condition: Non-obstructive Azoospermia
MeSH terms: Azoospermia
Study type: Interventional

Summary: 
      Infertility is a significant social- and health problem in the Western World and at the
      moment in Denmark one in ten babies are born with the help of assisted reproduction. In 50%
      of infertile couples a male factor can be identified as a contributing cause (1).

      Azoospermia is defined as the absence of spermatozoa in the ejaculate and it is a condition
      affecting 10-15% of infertile men (2, 3). Azoospermia is divided into obstructive azoospermia
      (OA) and nonobstructive azoospermia (NOA) of which the latter constitutes 60% (2, 3). In NOA
      the production of spermatozoa in the testis is either absent or markedly decreased.

      Since 1999 microdissection testicular sperm extraction (mTESE) has become the preferred
      treatment option for NOA in many centers worldwide (4). The procedure is performed in general
      anesthesia using an operating microscope to carefully examine the entire testicular tissue
      for the presence of spermatozoa which can be used for assisted reproduction.

      An alternative to mTESE is a percutaneous testicular sperm aspiration (TESA) or needle
      biopsy. This procedure is simple to perform using a biopsy needle to aspirate testicular
      tissue. The aspirated tissue is examined for the presence of spermatozoa that can be used in
      assisted reproduction.

      Today there is no robust evidence on the optimal sperm retrieval protocol on men with NOA.
      This is in part due to the fact that no randomized trials have been performed to compare
      procedures. This study is the first to randomize procedures for surgical sperm retrieval.

      Hypothesis In men with NOA, the investigators hypothesize that TESA is a viable first line
      approach compared to mTESE in regards to success rates of finding spermatozoa, complication
      rates and pregnancy outcomes.

      A total of 110 men will be randomized to either mTESE or TESA and the rates of finding
      spermatozoa will be compared. However, for ethical reasons, because some believe mTESE have a
      greater chance of finding sperm cells, all men with a failed TESA will have a mTESE
      afterwards.
    

Detailed description: 
      Background Infertility is a significant social- and health problem in the Western World and
      at the moment in Denmark one in ten babies are born with the help of assisted reproduction.
      In 50% of infertile couples a male factor can be identified as a contributing cause (1).

      Azoospermia is defined as the absence of spermatozoa in the ejaculate and it is a condition
      affecting 10-15% of infertile men (2, 3). Azoospermia is divided into obstructive azoospermia
      (OA) and nonobstructive azoospermia (NOA) of which the latter constitutes 60% (2, 3). In NOA
      the production of spermatozoa in the testis is either absent or markedly decreased.

      Since 1999 microdissection testicular sperm extraction (mTESE) has become the preferred
      treatment option for NOA in many centers worldwide (4). The procedure is performed in general
      anesthesia using an operating microscope to carefully examine the entire testicular tissue
      for the presence of spermatozoa which can be used for intracytoplasmic sperm injection
      (ICSI). The advantages of mTESE include high sperm retrieval rates (SRR), in recent
      systematic reviews reported around 52% (5-8), and low reported complication rates (9-11).
      However mTESE is a time-consuming invasive procedure requiring the right expertise, the right
      equipment and a proper setup of the fertility clinic.

      An alternative to mTESE is a percutaneous testicular sperm aspiration (TESA). This procedure
      is simple to perform using a biopsy needle to aspirate testicular tissue. The aspirated
      tissue is examined for the presence of spermatozoa and SRR for this procedure is usually
      reported around 25% (5-8). The lower SRR compared to mTESE is the main limitation of this
      method, but the procedure is well tolerated by patients and due to its simplicity and short
      duration it can easily be performed in the out-patient clinic. In collaboration with
      University of Michigan the TESA technique has been improved by using an 18 gauge needle and
      performing multiple (50-100) passes throughout the entire testicular tissue while applying a
      vacuum. In the remainder of this document this multiple needle-pass TESA will be referred to
      as "TESA". Recently the investigators retrospectively evaluated this approach and found a SRR
      of 30% using TESA and interestingly the SRR of mTESE following a failed TESA was only 11%
      (12).

      However, there is no robust evidence on the optimal sperm retrieval protocol on men with NOA.
      This is in part due to the fact that no randomized trials have been performed to compare the
      two procedures. Often patients from different countries are required to pay the infertility
      treatment themselves and the cost for a TESA is around $500 while a mTESE is around $8.000.
      This makes it difficult to perform a randomized trial in most countries but because of the
      public health systems of Denmark and Sweden there is a unique possibility to be the first in
      the world to do a randomized trial on this matter.

      Hypothesis In men with NOA, the investigators hypothesize that TESA is a viable first line
      approach compared to mTESE in regards to SRR, complication rates and pregnancy outcomes.

      Screening and randomization Potential study patients and their partners will initially be
      identified in their work-up during their infertility treatment. When a patient is identified
      as having NOA, the patient will be screened according to inclusion- and exclusion criteria.
      If these are met and the patient provides an informed consent for participation, the patient
      is included and randomized to undergo mTESE or TESA. Patients will be recruited from the
      fertility clinics at Herlev and Gentofte Hospital, Malmö University Hospital, Holbaek
      Hospital and Rigshospitalet in collaboration with Department of Growth and Reproduction,
      Rigshospitalet.

      Planning the treatment and surgery Treatment of the infertile couples will follow regular
      treatment protocols at the respective fertility clinics. This includes freezing retrieved
      sperm cells and thawing them in relation to the oocyte retrieval. As an alternative to sperm
      cell freezing, retrieved oocytes will be frozen and thawed in relation to the surgical sperm
      retrieval. The aim is to have 6-8 oocytes stored before operation for couples following this
      route. All patients will undergo full anesthesia. If the patient is randomized to TESA and
      this fails, then a salvage mTESE can be done immediately while in the operating theatre.

      Visits Before visit 1 all patients and their partners will have received written and oral
      information on the study and will have had time to provide an informed consent.

      At visit 1 patients and possibly their partners (mandatory in cases of oocyte
      cryopreservation) meet with the primary investigator. It will be secured that an informed
      consent for participation is signed. The focus of visit 1 will be on accessing the men's
      evaluation leading to the NOA diagnosis and checking inclusion- and exclusion criteria. In
      addition, information regarding medical history will be obtained and a physical examination
      of the external genitalia will be carried out. Patients will fill out questionnaires on
      psychosexual health and pain before the visit and their answers will be reviewed together. A
      blood sample will be taken for the purpose of measuring hormone levels and for storage in the
      biobank. Likewise patients will bring a semen sample for storage in the biobank. If not
      previously done patients will be tested with regards to karyotype, AzF microdeletions and
      CFTR mutations if deemed necessary.

      Visit 2 will be the actual procedure. Follow-up is comprised by patients filling out the
      mentioned questionnaires, reporting any complications and having a blood sample drawn at
      visit 3 and 4. Follow-up will primarily be done by phone/mail, but at visit 3 and 4 patients
      will have to go to the Department of Growth and Reproduction, Rigshospitalet (for Danish
      patients) or the fertility clinic at Malmö University Hospital (for Swedish patients) to have
      blood samples drawn. Samples collected in Sweden will be send for analysis at the Department
      of Growth and Reproduction, Rigshospitalet. Pregnancy outcome will be reported by phone/mail
      9-15 months after the procedure is done.

      Biobank A research biobank will be made containing blood samples, semen and testicular
      tissue. Blood samples will be used for genotyping and to measure relevant biomarkers
      including piRNAs, miRNAs, protein markers and biochemical markers. Semen will also be used to
      measure relevant biomarkers. Testicular tissue will be used for testing for molecular and
      biochemical markers of spermatogenesis and to measure intratesticular testosterone levels.
      Additionally testicular tissue showing maturation arrest will be used for attempts to
      maturate germ cells in vitro (not to be used for fertilization, only research). The purpose
      of the above is to find markers of successful sperm retrieval and to get a better
      understanding of the biology of NOA and potentially to develop pharmacological strategies in
      the future for treating certain forms of NOA as e.g. maturations arrest.

      Safety and adverse events:

      Hematoma As TESA/ mTESE is a surgery there is a risk of post-operative bleeding leading to
      scrotal or spermatic cord hematomas. Estimated risk <1,5%.

      Infection As TESA/ mTESE is a surgery there is a risk of infection. Estimated risk <1% Pain
      Pain is expected to occur after both procedures but especially after mTESE Drop in
      testosterone level In the litterature this is reported as a transient event following mTESE
      however there is a risk of more lasting androgen deficiency. Careful surgical techniques will
      be used to limit the amount of tissue damage.

      Possible disadvantages:

      Lower SRR following TESA SRR are reported lower for TESA compared to mTESE. For this reason
      patients with an unsuccessful TESA will be offered a salvage mTESE Oocyte cryopreservation In
      the past fertilization outcomes after cryopreservation have been lower compared to fresh
      oocytes. However with the evolution of cryopreservation techniques (vitrification) IVF
      pregnancy rates with the use of cryopreserved oocytes are now similar to those of fresh
      oocytes. (15) Grading of complications Complications will be graded according to the
      Clavien-Dindo classification of surgical complications (14).

      Ethical evaluation This study will follow the newest edition of the Helsinki Declaration and
      at all times follow applicable laws. It will be reported to "Datatilsynet", "Videnskabsetisk
      komite" and the "Ethical Committee of Lund University".

      Some of the ethical aspects related to this project have been briefly described in the safety
      and adverse events section. Both mTESE and TESA are safe procedures and although the
      investigators previously found a higher complication rate with mTESE compared to TESA (12),
      the general literature describe both procedures with comparable risks related to infection
      and hematomas.

      To make the study possible to complete, either the sperm cells or the oocytes retrieved from
      the women will have to undergo cryopreservation. Sperm cell cryopreservation is standard
      practice at the fertility clinics today. Previously only fresh oocytes were used for in-vitro
      fertilization but recently it has been shown that treatment using cryopreserved oocytes give
      similar pregnancy rates to those of fresh oocytes. Thus freezing the oocytes should not be a
      disadvantage to the couples.

      When looking at sperm retrieval rates, mTESE has until now been superior to TESA. However
      with the improved TESA results are expected to be more equal. In addition patients randomized
      to TESA will have the benefit of a more simple and less invasive treatment compared to mTESE.
      Moreover patients randomized to TESA will be provided with an option to receive a salvage
      mTESE that can be carried out the same day because all patients will undergo full anesthesia.
      In this way all patients included in the study will receive equal treatment options for NOA.

      Attempts of maturation of germ cells in vitro will be made using tissue with partial or total
      maturation arrest. If successful, the germ cells maturated in vitro will not be used for any
      attempt of fertilization but only as "proof of concept" regarding treatment of azoospermia
      due to maturation arrest.

      In a broader picture this project will be the first randomized study on this matter and
      therefore it will bring a clearer picture on which is the optimal sperm retrieval technique
      for men with NOA. This will benefit many patients with NOA in the future.

      Publication of results:

      Results from the study will be published in international peer-reviewed scientific journals,
      national journals and hospital websites. This will happen no matter the outcome of the study
      meaning that both positive, negative and inconclusive results will be made public.
      Additionally results will be presented at conferences and meetings. All published results
      will be made anonymous.
    

Intervention name(s): mTESE; TESA
Intervention type(s): Procedure; Procedure
Intervention description(s): mTESE is performed according to the initial description by Schlegel (4). General anesthesia is used. Midline incision of the scrotum is made and carried down to the level of the tunica vaginalis, which is opened to deliver the testis. Equatorial incision is made and the testis bivalved. The operating microscope is used to identify plump-appearing seminiferous tubules, and biopsies are taken from these areas. These samples are examined by an embryologist in the operating room after initial gross dispersion by passing the tissue through a 24G angiocatheter. The procedure is stopped if spermatozoa are found. If no plump tubules are found, random biopsies targeting every area of the testis are taken. If necessary the procedure is carried out on the contralateral testis. Hemostasis is obtained with careful bipolar cautery, and the tunica albuginea and the tunica vaginalis are closed with a running 4-0 Vicryl. Standard scrotal closure is performed and gentle pressure dressing is applied.; In general anesthesia an 18 gauge needle is introduced into each testis, and negative pressure applied with a 10 ml syringe. Multiple passes throughout the entire testis, numbering 50-100 passes, are made through a single percutaneous/tunical entry and continued until tissue is visible in the hub of the needle. This is removed by brisk extraction of the needle and pressure held to tamponade bleeding. The aspirated tissue is examined by an embryologist in the operating room after initial gross dispersion by passing the tissue through a 24 gauge angiocatheter.
Other intervention name(s): Microdissection testicular sperm extraction; Percutaneous testicular sperm aspiration
Allocation: Randomized
Intervention model: Parallel Assignment
Intervention model description: Trial type:
Clinical randomized multi-center controlled trial
Randomization:
Block randomization is performed with the use of a computer generated block randomization list
Participants:
Patients with NOA undergoing fertility treatment
Masking: Single (Participant)
Primary purpose: Treatment
Keywords: azoospermia; microdissection testicular sperm extraction; testicular sperm aspiration; male infertility; surgical sperm retrieval