Neoplasms

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Thyroid Diseases

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Iodine

Cadexomer iodine

Fresh semen treated without irradiation before cryopreservation

cryopreserved semen without irradiation (n=60)

Pelvis scanning : acute irradiation (1 or 2 second) using low dose (n=30),

Bone scintigraphy : long irradiation (3h) using low dose (n=30),

Radioactive Iodine 131 therapy (RAT) applied for thyroid cancer : long irradiation (3h) using medium dose (n=60),

sperm will be frozen in Cryosperm® cryoprotectant medium (Origio). The samples will be packaged in previously identified high security straws (Cryobiosystem). Slow freezing of the straws will be carried out using the Nanodigicool® programmable device (Cryobiosystem).

Cryopreserved semen

sperm sample will be placed on the scanner's processing table to be exposed for 1 to 2 seconds. Several dose levels will then be made in order to limit the value of 17 mGy

Acute and low irradiation

sperm sample will be exposed to gamma radiation by adding a solution of Tc99m for 3 hours.

Long and low irradiation

sperm sample will be exposed to gamma radiation by adding a solution of I131 for 3 hours

Long and medium irradiation

Lise Laclautre, PharmD

Hanaé PONS-REJRAJI, PhD

Florence BRUGNON, MD, PhD

Solène VORILHON, MD, PhD

Cyril BOUCHE, PhD

Aurélie VEGA, PhD

Philippe VAGO, MD, PhD

Andreï TCHIRKOV, MD, PhD

Céline PEBREL-RICHARD, MD

Gaëlle SALAUN, MD, PhD

Lauren VERONESE, MD, PhD

Laetitia GOUAS, MD, PhD

Carole GOUMY, MD, PhD

Hanae PONS-REJRAJI, PhD

Differentiated thyroid cancer is the third cause of cancer in young men of childbearing age. Its treatment by irradiation with Radioactive Iodine 131 therapy (RAT) could alter spermatogenesis and result in azoospermia and permanent infertility. A preventive gametes cryopreservation was recommended before RAT, but without mentioning a period of teratogenic risk transmissible to the offspring. To date, RAT impact on human sperm nucleus is poorly known or even unknown, notably on telomere length.

Our objective is to define RAT effects on human sperm nucleus by in vitro irradiation exposure of human spermatozoa to mimicking that of the gonads in the context of irradiation with iodine131 used for thyroid cancer. We will analyze standard sperm parameters, major DNA alterations and telomere length using molecular and cellular assays. Nucleus morphology and chromatin organization will also be analyzed using 3D bio-imaging. This study will permit to optimize the indications for the preservation of fertility.

Our main objective is to measure the in vitro impact of irradiation treatment on sperm nuclear quality such as DNA fragmentation and oxidation, chromatin condensation and organisation, nucleus morphology and notably sperm telomere length (STL).

The secondary objectives are :

* to measure RAT impact on sperm parameters (vitality, motility and morphology)
* to measure the impact of cryopreservation on chromatin organisation and nucleus morphology
* to evaluate RAT impact on human sperm cells in comparison with cryopreservation
* to evaluate the impact of different doses and types of irradiation on human sperm cells
* to establish relations between potential alterations of standards sperm parameters (vitality, motility and morphology) and nuclear sperm parameters (DNA fragmentation and oxidation, chromatin condensation and organisation, nucleus morphology and STL.

Our final goal is to provide a significant improvement of men fertility diagnosis and optimize our fertility preservation practices.

To this end, we will expose ejaculated human spermatozoa (n = 90) at different (low and moderate) doses of gamma or X photons to mimic gonads irradiation. Absorbed doses by the samples were calculated using the GATE Monte Carlo platform (version 8.2). Experiment geometry settings were modelled as three dimension voxelized volumes inside the software by assigning a shape, size, distance and density for all the volumes created. All measurements will be made on surplus samples from men undergoing routine semen analysis at the Center for Reproductive Medicine, after receiving their written informed consent. Semen samples are collected and subdivided into 3 arms to analyse sperm quality after:

* irradiation exposure (3 conditions);
* a freezing- thawing cycle;
* fresh state: negative control without treatment.

Before (fresh state) and after each treatment we will analyse:

* standard semen parameters (vitality, motility and morphology) in accordance with WHO, 2010;
* STL using Flow FISH, q-PCR and q-FISH;
* chromatin condensation (chromomycin A3);
* DNA oxidization (8-OHdG residues);
* DNA fragmentation (TUNEL);
* 3D nucleus structure using 3D bio-imaging.

Impact of Ionizing Treatment on the Nuclear Structure of Human Spermatozoa.

To achieve this goal, we will expose human sperm to an irradiation dose of I131 to mimic the irradiation of gonads and sperm. Reliquat of semen will be cryopreserved (control condition). After thawing, the same sample will be subdivised into 2 groups in order to analyze the sperm quality :

* After an irradiation dose of I131
* After a cryopreservation

All measurements are made on surplus samples from men undergoing routine semen analysis at the Center for Reproductive Medicine, after receiving their written informed consent. Semen samples will be collected and subdivided into 3 arms to analyse sperm quality after:

* irradiation exposure (3 conditions);
* a freezing- thawing cycle;
* fresh state: negative control without treatment.

Before (fresh state) and after each treatment we will analyse:

* STL using Flow FISH, q-PCR and q-FISH;
* chromatin condensation (chromomycin A3);
* DNA oxidization (8-OHdG residues);
* DNA fragmentation (TUNEL);
* 3D nucleus structure using 3D bio-imaging.

All measurements are made on surplus samples from men undergoing routine semen analysis at the Center for Reproductive Medicine, after receiving their written informed consent. Semen samples will be collected and subdivided into 3 arms to analyse sperm quality after:

* irradiation exposure (3 conditions);
* a freezing- thawing cycle;
* fresh state: negative control without treatment.
* a freezing- thawing cycle;
* fresh state: negative control without treatment. Before (fresh state) and after each treatment we will analyse standard semen parameters (vitality, motility and morphology) in accordance with WHO, 2010;

We will compare effects induced by RAT and cryopreservation on each sperm sample. In order to limit the bias, for each irradiation treatment, a sample of sperm cells (cryopreserved control) will be treated under identical conditions.

All measurements are made on surplus samples from men undergoing routine semen analysis at the Center for Reproductive Medicine, after receiving their written informed consent. Semen samples will be collected and subdivided into 3 arms to analyse sperm quality after:

* irradiation exposure (3 conditions);
* a freezing- thawing cycle;
* fresh state: negative control without treatment.

Irradiation exposure will be realised under 3 conditions:

* acute and low irradiation : sperm sample will be placed on the scanner's processing table to be exposed for 1 to 2 seconds. Several dose levels will then be made in order to limit the value of 17 mGy.
* long and low irradiation: sperm sample will be exposed to gamma radiation by adding a solution of Tc99m for 3 hours.
* long and medium irradiation : sperm sample will be exposed to gamma radiation by adding a solution of I131 for 3 hours

We will correlate the effects induced by on vitality, motility and morphology and nuclear sperm parameters (DNA fragmentation and oxidation, chromatin condensation and organisation, nucleus morphology and STL.

Study title	NCT ID	Conditions	Interventions	Eligibility	Organization	Link
Impact of Ionizing Treatment on the Nuclear Structure of Human Spermatozoa.	NCT04715828	Sperm Preservat... Irradiated Seme...	Cryopreserved s... Acute and low i... Long and low ir... Long and medium...	18 Years - 45 Years	University Hospital, Clermont-Ferrand
Impact of Ionizing Treatment on the Nuclear Structure of Human Spermatozoa.	NCT04715828	Sperm Preservat... Irradiated Seme...	Cryopreserved s... Acute and low i... Long and low ir... Long and medium...	18 Years - 45 Years	University Hospital, Clermont-Ferrand

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