Résumé: Lorsqu’une femme en âge de reproduction est diagnostiquée avec un cancer du sein, le traitement comprend entre autres de la chimiothérapie et/ou une hormonothérapie. Malheureusement, certaines patientes deviendront infertiles suite à la chimiothérapie qui est gonadotoxique, ce qui diminue leurs chances de devenir mères avec leurs propres gamètes. Etant donné que la grossesse après un cancer du sein n’est plus considérée comme un facteur de risque de récidive de la maladie, l’accès à la maternité chez ces jeunes patientes est devenu un enjeu majeur. Cependant, peu de données ont été publiées concernant la sécurité de concevoir grâce à la PMA chez une patiente devenue infertile suite au traitement du cancer du sein.Il est donc primordial d’offrir aux femmes qui le désirent une méthode de préservation de la fertilité avant tout traitement gonadotoxique.La conservation d’ovocytes ou d’embryons après hyperstimulation ovarienne est une des techniques offertes aux patientes. Néanmoins, en cas de maladie hormono- sensible tel que le cancer du sein, une élévation d’hormones stéroïdiennes accompagnant la réponse ovarienne est peu souhaitable. Des protocoles modifiés ont donc été développés, tel que l’hyperstimulation ovarienne aux gonadotrophines associée au letrozole, un inhibiteur de l’aromatase.Malgré certains résultats rassurants dans la littérature, les données concernant l’efficacité et la sécurité de ce protocole sont toutefois limitées.Notre projet comporte 2 objectifs:1. Evaluation d’une procédure de préservation de la fertilité associant une hyperstimulation ovarienne au letrozole (Let-COH), dans une étude prospective non randomisée (BROVALE).2. Evaluation de la sécurité de la grossesse associée à la PMA chez des patientes ayant été traitées pour un cancer du sein.Dans la première partie nous avons confirmé l’efficacité du Let-COH sur base du taux de maturation ovocytaire similaire à une population contrôle de patientes7infertiles ayant bénéficié d’une hyperstimulation ovarienne conventionnelle sans letrozole (groupe contrôle) quel que soient les modes de déclenchement de la maturation ovocytaires utilisés, human chorionic gonadotropin (hCG) ou l’agoniste de la gonadotropin-releasing hormone (GnRHa). Par ailleurs, les modifications endocriniennes dans le liquide folliculaire, et l’expression de gènes liés à la qualité ovocytaire au niveau des cellules du cumulus oophorus (CC) ont été analysées. Dans le microenvironnement entourant l’ovocyte, nous avons observé un taux d’œstradiol significativement plus bas et un taux de testostérone significativement plus élevé dans le groupe Let-COH, comparé au groupe contrôle. Par ailleurs, l’expression génique au niveau des CC était plus basse dans le groupe Let-COH déclenché à l’hCG comparé au groupe contrôle. Cependant l’inverse était observé lorsque le GnRHa était utilisé comme déclencheur de l’ovulation. Ces résultats suggèrent donc un effet bénéfique des analogues de la GnRH sur la qualité ovocytaire avec le Let-COH.Nous avons montré que le taux d’oestradiol était bas durant la stimulation, mais que le taux de progestérone durant la phase lutéale était supra-physiologique chez les patientes ayant reçu le Let-COH, comparable au taux des patientes contrôles.Etant donné le rôle potentiel de la progestérone dans la carcinogenèse mammaire, nous avons modifié le protocole de déclenchement de l’ovulation, pour administrer le GnRHa au lieu de l’hCG.Dans la seconde partie de notre travail, nous avons réalisé une étude rétrospective multicentrique afin de comparer les issues oncologiques et de grossesse de patientes ayant été enceintes après cancer du sein, naturellement (n=173) ou grâce à la PMA (n=25). Malgré la petite taille de l’échantillon obtenu, nous n’avons pas observé plus de récidives chez les patientes ayant eu recours à la PMA.En conclusion, ce projet nous a permis de démontrer l’efficacité de la préservation de la fertilité chez les jeunes patientes atteintes de cancer du sein, par hyperstimulation associée au letrozole. Nous avons également observé que le risque de récidive de la maladie n’était pas majoré dans notre échantillon limité de patientes enceintes suite à la PMA après cancer du sein. / Abstract:Young women diagnosed with breast cancer are often treated with adjuvant primary therapy, including chemo and/or endocrine therapy. These women may face infertility or premature ovarian failure due to the gonadotoxicity of chemotherapy regimens or to aging, which significantly decreases their chances to become mothers with their own gametes. Since it has been established that pregnancy after breast cancer does not increase the risk of relapse, assisting young breast cancer survivors to access motherhood has become a major quality of life issue. However, data on the safety of pregnancies associated with ART in patients who became infertile are scarce. Therefore, it is recommended that young patients diagnosed with breast cancer be informed about treatment-related infertility risks and available procedures to preserve their fertility before undergoing gonadotoxic treatment.Oocyte and/or embryo cryopreservation following controlled ovarian hyperstimulation (COH) is the most established fertility preservation procedure that patients can undergo before initiation of chemotherapy. However, this procedure induces an increase in steroid levels, which may be detrimental in hormonally sensitive diseases such as breast cancer. In order to avoid this issue, a modified protocol has been developed a decade ago, combining letrozole (an aromatase inhibitor) with conventional COH (Let-COH).Despite recent reassuring preliminary results reported in the literature, data are still very limited regarding the efficiency and the safety of this protocol.Thus, the aims of this project were:1. Evaluation of the efficiency of Let-COH before adjuvant therapy, in anonrandomized prospective study (BROVALE).2. Evaluation of the safety of ART-associated pregnancy in breast cancersurvivors.In the first part, we confirmed the efficiency of Let-COH (study group) based on similar oocyte maturation rates when compared to infertile patients undergoing conventional COH without letrozole (control group), regardless of the triggering5method, human chorionic gonadotropin (hCG) or GnRH-agonist (GnRHa). Oocyte quality was indirectly assessed by analysis of follicular fluid (FF) steroid levels and cumulus cell (CC) gene expression (HAS2, PTGS2, and GREM1). We found that estradiol levels were significantly lower and testosterone levels significantly higher in the study compared to the control group, suggesting lower oocyte quality. Nevertheless, when GnRHa was used as ovulation trigger, differences in estradiol levels between the groups were reduced. Similarly, CC gene expression was lower in the study group compared to the control group in the hCG triggered subpopulation, while the opposite effect was observed in the GnRH-agonist triggered subpopulation. Altogether, the results suggest a benefit of GnRHa trigger on oocyte quality in Let- COH protocol.The safety of the protocol was assessed by measuring estradiol and progesterone levels during and after Let-COH. We confirmed that estradiol levels remained low during treatment. However, during luteal phase, progesterone levels were comparable to conventional COH when triggered with hCG, but not with GnRHa. Since progesterone may be as important as estradiol in promoting breast carcinogenesis, we modified the protocol to systematically use GnRH-agonist as ovulation trigger.In the second part of our project, we conducted a retrospective multicenter study to compare pregnancy and oncological outcomes among breast cancer survivors who subsequently conceived spontaneously (n=173) or following ART (n=25). Although the ART group’s sample size was small, we did not observe an increased relapse rate after ART compared to patients who conceived naturally.In conclusion, we confirmed the feasibility and efficiency of Let-COH as a method of fertility preservation in breast cancer patients. Moreover, in our small series of ART- associated pregnancies in breast cancer survivors we did not observe an increased risk of disease recurrence. / Doctorat en Sciences médicales (Médecine) / info:eu-repo/semantics/nonPublished

Cryopreservation of gametes and embryos is crucial in fertility treatment and fertility preservation. Preservation of oocytes is a more complicated process than preservation of sperm and embryos. According to recent studies, a new preservation technique called vitrification is found to have higher rates of oocytes-survival after warming    The aim of this study was to evaluate VitriBlast™ Kit, for vitrification of oocytes. Vitrification is an ultra-rapid freezing method in the presence with of high concentrations of cryoprotectants which avoids intracellular ice-crystal formation during the freezing and warming process.     In this study, a total number of 117 immature oocytes were used and 62 of these oocytes were vitrified with VitroBlast™ Kit. During this process two different vitrification devices were used, VitroLoop™ and Cryopette®. After warming, the average survival rate for vitrified oocytes was found to be 61% for VitroLoop™ and 15% for Cryopette® (p<0.001). The remaining 55 oocytes were used as a non-frozen control group and the same incubation method as for vitrified oocytes was used. The survival rate for the control groups was higher than for the vitrified groups (93% versus 35%, p<0.001). The results of this study indicate that VitriBlast™ Kit is not suitable for vitrification of oocytes.

BACKGROUND: Gliomas are the most common primary brain malignancy, with more than 16,000 patients diagnosed every year (Ostrom, et al., 2015). Outcomes vary widely depending on tumor grade and treatment, and have been steadily improving with the advent of new therapeutics. Glioma patients frequently undergo chemotherapy to remove residual tumor after surgery, and many of these cytotoxic therapies are known to affect rapidly dividing cells such as ovarian follicles (Vassilakopoulou et al., 2016). The negative effects of chemotherapy on fertility have been demonstrated in patients with breast and colorectal cancer (Bines, et al., 1996; Avastin Prescribing Information). Additionally, infertility has been linked with decreased quality of life, primarily in women (O’Moore et al., 1983; Greil, 1997). Fertility treatments are available for women undergoing cancer treatment, however it is unknown whether these treatments are routinely discussed with glioma patients before initiating chemotherapy. OBJECTIVE: The primary goal of this study was to assess whether female glioma patients are being effectively counselled on their possible loss of fertility and their choices for fertility treatment prior to beginning chemotherapy. To this end, it was also important to understand the barriers preventing patients from obtaining information related to their fertility. Another principle goal of this study was to describe the effects of chemotherapy on a sample of women with glioma. Finally, this study sought to understand the priorities of women with glioma in regards to family planning, and to address these priorities in the context of a comprehensive fertility preservation discussion. METHODS: To assess these endpoints, a survey was designed and delivered to patients being treated at the Neuro-oncology clinic of the University of California, San Francisco. Eligible candidates were identified prior to a clinic visit, and patients were asked whether they would like to participate in the survey. Consenting patients then completed the survey at home or in the clinic. Seventy two women completed the survey. Data was analyzed using STATA Software Version 10.0. RESULTS: Analysis of the survey results showed that only 35% of women receiving chemotherapy reported having a discussion regarding fertility preservation prior to beginning treatment. Of those who reported having this discussion, only 80% were aware that chemotherapy could negatively affect their fertility. Many women reported that while fertility preservation was not important to them at the time of diagnosis, it was a priority for them at the time of survey completion. Most women surveyed expressed a desire to have a fertility preservation discussion with a reproductive specialist. CONCLUSIONS: The data obtained in this study suggest a lack of understanding of the negative effects of chemotherapy which may be addressed with a more comprehensive fertility discussion with glioma patients prior to beginning treatment. Although interest in having children tends to decrease after cancer treatment, the majority of respondents still report wanting a child after treatment. The priorities of women in the study reflect a concern for the health of their future offspring which may be best addressed prior to beginning treatment in order to increase their chances of conceiving at a later date.

There is an increased number of childhood cancer survivors living into adulthood. As more survivors live into adulthood, researchers have been able to study and better understand the late effects of cancer treatment. A well-known late effect of cancer treatment is the risk of infertility. Cancer-related infertility is a source of distress to cancer survivors. There have been many advances to fertility preservation over the last few years and there are now multiple options available for both men and women. Despite the improved understanding of the risk of cancer-related infertility and advances to fertility preservation treatment, these services remain underutilized by cancer patients. It is known that discussing fertility preservation options with newly diagnosed cancer patients improves survivors’ long-term quality of life and reduces decisional regret, regardless of if they pursue fertility preservation treatment. Survivors often report that the risk of treatment-related infertility and/or available fertility preservation options was often inadequately or not discussed with them at the time of diagnosis. The use of fertility preservation decision aids for adult patients newly diagnosed with cancer have been proven to be effective at improving participants’ knowledge surrounding fertility preservation, reducing decisional conflict, and reducing long term decisional regret. A fertility preservation decision aid has not yet been developed for use by adolescents newly diagnosed with childhood cancer. This study aims to engage survivors and providers to develop a fertility preservation decision aid to improve the decision quality of adolescents newly diagnosed with childhood cancer who are determining their preferences on accepting a referral to a fertility specialist. This study then proposes to field test the decision aid with newly diagnosed patients. The use of a decision aid will lead adolescents with childhood cancer to have increased knowledge on the risk of infertility and the fertility preservation options available. This study also aims to lower participants’ levels of decisional conflict about their fertility preferences. There is a need to incorporate the use of a fertility preservation decision aid into childhood cancer treatment. If this decision aid proves effective, referral to the fertility preservation decision aid may become common practice at the time of initial diagnosis. If the decision aid is effect at improving decision quality and reducing decisional conflict, survivors may experience long-term benefits including improved quality of life and reduced levels of decisional regret.

BACKGROUND: There are currently an estimated 250,000 female cancer survivors of reproductive age living in the US. Loss of fertility is an issue many cancer survivors face after treatment, as all forms of cancer therapy can cause infertility. Methods to preserve fertility can be initiated prior to cancer therapy. These methods include embryo cryopreservation, oocyte cryopreservation, fertility sparing surgery, ovarian tissue cryopreservation, ovarian transposition, and medical therapy. LITERATURE REVIEW: Although the clinical guidelines state that oncologists should discuss the risk of infertility with every patient of reproductive age and should refer every patient who is interested in or ambivalent towards fertility preservation to reproductive endocrinologists, studies have shown that a significant proportion of female cancer patients report never receiving information about fertility. Even fewer female cancer patients are referred to reproductive endocrinologists for further discussion and/or potential treatment. PROPOSED PROJECT: Oncologists at Boston Medical Center will be recruited to participate in a study that measures the effect of an educational intervention on referral rate to reproductive endocrinology. The knowledge gained from the intervention will be assessed with a pre- and post-test. The proportion of female patients age 18-45 referred to reproductive endocrinology will be evaluated through the Electronic Medical Record System. The correlation between knowledge gain and change in referral rates will also be assessed. CONCLUSION: Fertility after cancer treatment is an essential issue to consider for young cancer survivors. These patients benefit from being referred to reproductive endocrinologists so that they can get information about fertility preservation and undergo treatment in a timely fashion. Improving and/or reinforcing oncologist knowledge about this topic will increase the rate at which they initiate this conversation and therefore the number of female patients who are referred to reproductive endocrinology. SIGNIFICANCE: Providing female cancer patients with information about and opportunities to undergo fertility preservation will maximize their options. This will lead to a higher quality of life after cancer therapy.

Adolescents and young adults (AYAs) ages 15-39 with cancer have experienced stagnant survival rates for the past 30 years in comparison to those older or younger diagnosed with cancer. Survival disparities for this population may be due to biologic factors, lack of consistent and effective access to care, and unique psychosocial needs of this age group, and taken together, present an opportunity for health policy intervention. Of particular interest are barriers most important to AYAs themselves, appropriate and timely clinical trial enrollment, and the ability to preserve fertility prior to initiating treatment. These barriers may be more difficult to overcome for AYAs that are from rural areas, those that may be un- or under- insured, of lower socioeconomic status or educational attainment, and other social determinant of health related-factors. In this dissertation, I examined the factors that drive the challenges encountered by AYAs with cancer and discuss potential solutions to overcome these challenges. The first aim of this dissertation establishes which challenges or aspects of the cancer experience are most important to AYAs using a mixed methods approach. The second and third aims build off of challenges identified in aim one using quantitative methods. In aim two, AYA clinical trial enrollment in the United States is examined using a nationally-representative sample with a specific focus on rural disparities. In aim three, historical utilization of fertility preservation consultations and procedures are studied among AYAs treated a single institution serving a rural population.

Orientador: Anaglória Pontes / Resumo: O câncer de mama é uma doença maligna relativamente comum em mulheres adultas jovens, justificando a preocupação com a potencial toxicidade gonadal relacionada à quimioterapia. É importante considerar o encaminhamento precoce de pacientes jovens com câncer de mama que tenham desejos reprodutivos para especialistas, a fim de discutir sobre a preservação da fertilidade. A criopreservação de embriões ou de oócitos estão entre os principais métodos à disposição, e para conseguir isso, estimulação ovariana controlada (EOC) é o primeiro passo a ser considerado. O presente estudo tem como objetivo apresentar protocolo de estimulação ovariana em para preservação da fertilidade em pacientes com câncer de mama. De novembro de 2014 a dezembro de 2016, 109 pacientes com câncer de mama com menos de 40 anos de idade foram selecionadas para preservar seu potencial reprodutivo. Elas foram divididas de acordo com a fase do ciclo menstrual em que o estímulo ovariano foi iniciado: fase folicular inicial, fase folicular tardia e fase lútea. Para reduzir o tempo necessário da obtenção dos oócitos, este estudo utilizou o princípio do início aleatório, no qual a EOC pode ser iniciada durante qualquer período do ciclo menstrual sem consequências negativas. O letrozol foi utilizado durante toda a estimulação para diminuir as concentrações de estradiol, independentemente da imunohistoquímica tumoral. Na presença de tumores positivos ao estradiol, e indicação de quimioterapia neoadjuvante, o tamoxi... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: Breast cancer is a relatively common malignancy in young adult women, justifying the concern about the potential gonadal toxicity related to chemotherapy. It is important to consider early referral of young breast cancer patients with reproductive desires to fertility specialists, in order to discuss the fertility preservation. Embryos or oocytes cryopreservation are among the main methods available, and to achieve that, controlled ovarian stimulation (COS) is the first step to be considered. From November 2014 to December 2016, 109 breast cancers patients under 40 years were enrolled to preserve their reproductive potential. They were divided according to the menstrual cycle status in: initial follicular phase, late follicular phase and luteal phase. In order to reduce the time necessary to obtain the oocytes, this study used the principle of random start, in which the COS can be initiated during any period of the menstrual cycle without negative consequences. Letrozole was used during all stimulation to reduce estradiol concentrations, regardless of tumor immunohistochemistry. In the presence of estradiol positive tumors, and indication of neoadjuvant chemotherapy, tamoxifen was administerd as an additional protective measure. A GnRH agonist was used to trigger ovulation, in order to mitigate the risk of ovarian hyperstimulation syndrome. The following parameters were analyzed: age, day of COS start, number of days required to COS, total FSH dosage, estradiol levels at fi... (Complete abstract click electronic access below) / Mestre

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Agradecemos a compreensão. on 2017-06-30T12:25:51Z (GMT) / Submitted by FELIPE ANDREOTTA CAVAGNA null (cavfelipe@hotmail.com) on 2017-07-03T18:22:57Z No. of bitstreams: 1 Tese completa com ficha catalográfica.pdf: 2297427 bytes, checksum: 8e382c810380d26d332762e12fa2ab64 (MD5) / Approved for entry into archive by Luiz Galeffi (luizgaleffi@gmail.com) on 2017-07-04T16:36:48Z (GMT) No. of bitstreams: 1 cavagna_fa_me_bot.pdf: 2297427 bytes, checksum: 8e382c810380d26d332762e12fa2ab64 (MD5) / Made available in DSpace on 2017-07-04T16:36:48Z (GMT). No. of bitstreams: 1 cavagna_fa_me_bot.pdf: 2297427 bytes, checksum: 8e382c810380d26d332762e12fa2ab64 (MD5) Previous issue date: 2017-04-07 / O câncer de mama é uma doença maligna relativamente comum em mulheres adultas jovens, justificando a preocupação com a potencial toxicidade gonadal relacionada à quimioterapia. É importante considerar o encaminhamento precoce de pacientes jovens com câncer de mama que tenham desejos reprodutivos para especialistas, a fim de discutir sobre a preservação da fertilidade. A criopreservação de embriões ou de oócitos estão entre os principais métodos à disposição, e para conseguir isso, estimulação ovariana controlada (EOC) é o primeiro passo a ser considerado. O presente estudo tem como objetivo apresentar protocolo de estimulação ovariana em para preservação da fertilidade em pacientes com câncer de mama. De novembro de 2014 a dezembro de 2016, 109 pacientes com câncer de mama com menos de 40 anos de idade foram selecionadas para preservar seu potencial reprodutivo. Elas foram divididas de acordo com a fase do ciclo menstrual em que o estímulo ovariano foi iniciado: fase folicular inicial, fase folicular tardia e fase lútea. Para reduzir o tempo necessário da obtenção dos oócitos, este estudo utilizou o princípio do início aleatório, no qual a EOC pode ser iniciada durante qualquer período do ciclo menstrual sem consequências negativas. O letrozol foi utilizado durante toda a estimulação para diminuir as concentrações de estradiol, independentemente da imunohistoquímica tumoral. Na presença de tumores positivos ao estradiol, e indicação de quimioterapia neoadjuvante, o tamoxifeno foi utilizado como uma medida de proteção adicional. Um agonista do GnRH foi utilizado para desencadear a maturação folicular final, diminuindo o risco de síndrome de hiperestimulação ovariana. Foram analisados os seguintes parâmetros: idade das pacientes, dia de início da EOC, número de dias de EOC, dose total de FSH, níveis de estradiol no dia de maturação folicular final, número de oócitos coletados e número de oócitos vitrificados. A idade média dos pacientes foi de 31,27 ± 4,22 anos. A duração média da EOC foi de 10,0 ± 1,39 dias. O número médio de oócitos coletados foi de 11,62 ± 7,96 e o número médio de oócitos vitrificados foi de 9,60 ± 6,87. A concentração média de estradiol no dia do desencadeamento da ovulação foi de 706,30 ± 450,48 pg / mL, e a dose média de FSH administrada foi 2610,00 ± 716,51 UI. Ao comparar os resultados de acordo com a fase do ciclo no qual a EOC foi iniciada, não houve diferenças significativas no número de oócitos colhidos e vitrificados, duração da estimulação ovariana e dos níveis de estradiol no dia do desencadeamento da ovulação. Observou-se uma diminuição estatisticamente significativa da dose total de FSH administrada nos grupos que iniciaram a EOC na fase folicular tardia e na fase lútea, quando comparada com a fase folicular inicial. Esses resultados sugerem que a criopreservação de oócitos ou embriões com um protocolo específico para pacientes com câncer de mama, é eficaz e pode ser oferecida a mulheres jovens submetidas a tratamento citotóxico que têm preocupações relacionadas ao seu futuro reprodutivo. / Breast cancer is a relatively common malignancy in young adult women, justifying the concern about the potential gonadal toxicity related to chemotherapy. It is important to consider early referral of young breast cancer patients with reproductive desires to fertility specialists, in order to discuss the fertility preservation. Embryos or oocytes cryopreservation are among the main methods available, and to achieve that, controlled ovarian stimulation (COS) is the first step to be considered. From November 2014 to December 2016, 109 breast cancers patients under 40 years were enrolled to preserve their reproductive potential. They were divided according to the menstrual cycle status in: initial follicular phase, late follicular phase and luteal phase. In order to reduce the time necessary to obtain the oocytes, this study used the principle of random start, in which the COS can be initiated during any period of the menstrual cycle without negative consequences. Letrozole was used during all stimulation to reduce estradiol concentrations, regardless of tumor immunohistochemistry. In the presence of estradiol positive tumors, and indication of neoadjuvant chemotherapy, tamoxifen was administerd as an additional protective measure. A GnRH agonist was used to trigger ovulation, in order to mitigate the risk of ovarian hyperstimulation syndrome. The following parameters were analyzed: age, day of COS start, number of days required to COS, total FSH dosage, estradiol levels at final follicular maturation day, number of collected oocytes and number of vitrified oocytes. The mean age of patients was 31.27±4.22 years. The average duration of COS was 10.0±1.39 days. The mean number of collected oocytes was 11.62±7.96 and the mean number of vitrified oocytes was 9.60±6.87. The mean estradiol concentration on the day of the trigger was 706.30±450.48 pg/mL, and the mean dose of FSH administered was 2610.00±716.51 IU. When comparing the outcomes according to the phase of the cycle in which COS was commenced, there were no significant differences in the number of oocytes collected and vitrified, ovarian stimulation length and estradiol levels on the day of the trigger. It was observed a statistically decrease of the total FSH dose administered in the groups starting COS in the late follicular phase and in the luteal phase, when compared to the initial follicular phase These results suggest that oocyte or embryos cryopreservation with a specific protocol for breast cancer patients, is effective, safe, and may be offered to young women undergoing cytotoxic treatment who have concerns related to their reproductive future.

De nombreux agents chimiothérapeutiques sont gonadotoxiques et peuvent donc induire une insuffisance ovarienne précoce chez les jeunes patientes traitées. La protection pharmacologique de l'ovaire pendant la chimiothérapie à l'aide d'analogues de la Gonadotropin-Releasing Hormone (GnRHa) est une option intéressante de préservation de la fertilité de par son caractère non-invasif et la possibilité d’une récupération spontanée de la fonction ovarienne. Ces molécules sont des inhibiteurs bien connus de l'axe hypothalamo-hypophyso-gonadique, mais leur efficacité dans cette indication est, cependant, controversée et leurs mécanismes d'action sont mal compris. Par conséquent, nous avons investigué les mécanismes potentiels de protection ovarienne des GnRHa pendant la chimiothérapie sur modèle murin. Nous avons montré que le cyclophosphamide (Cy) induit une déplétion folliculaire aiguë et proportionnelle à la dose affectant à la fois les follicules quiescents et en croissance. Lorsqu'ils sont administrés seuls à différentes doses et sites, l'agoniste et l'antagoniste de la GnRH altèrent les cycles oestraux, mais ne bloquent ni la folliculogenèse ni la sécrétion de la Follicle-Stimulating Hormone (FSH) chez la souris. De plus, le Cy atteint les follicules primordiaux, que les souris aient été traitées avec les GnRHa ou non. Ces résultats suggèrent que les GnRHa n'inhibent pas l'axe hypophyso-gonadique aussi efficacement chez la souris que chez la femme. Par conséquent, nous avons développé de nouveaux modèles pour étudier les mécanismes potentiels de protection ovarienne des GnRHa. Afin de différencier les effets directs des GnRHa via leurs récepteurs ovariens ou indirects par inhibition de la sécrétion de gonadotrophines, l'effet de l'agent alkylant sur le développement folliculaire et la réserve ovarienne a été testé sur des follicules cultivés in vitro avec ou sans GnRHa et in vivo chez des souris déficientes en FSHb (Fshb-/-). Pour imiter la profonde inhibition de FSH observée chez la femme après traitement aux GnRHa, nous avons étudié la toxicité de la chimiothérapie chez les souris Fshb-/-. L’administration de gonadotrophines exogènes (pregnant mare serum gonadotropin, PMSG) induit une croissance folliculaire jusqu’au stade antral mais n’influence pas le nombre total de follicules au sein de l’ovaire. Le Cy induit une perte folliculaire significative dans le groupe contrôle et dans le groupe traité au PMSG. Aucune différence concernant la prolifération ni l'apoptose n'a été observée entre les groupes traités à la chimiothérapie. A ce jour, ce modèle murin représente le meilleur modèle pour étudier l'inhibition gonadotrope induite par les GnRHa observée chez la femme. Ces résultats suggèrent que la FSH n'est pas impliquée dans la protection ovarienne potentielle des GnRHa pendant la chimiothérapie. Afin d’évaluer les effets directs des GnRHa sur les follicules en croissance et quiescents, des follicules préantraux ou des ovaires de nouveau-nés (PND4) ont été cultivés avec ou sans GnRHa avant l'exposition au métabolite actif du Cy, le 4-hydroperoxycyclophosphamide (4HC). Nous avons d'abord montré que l'exposition in vitro aux GnRHa n'affectait ni la survie et le développement folliculaire, ni la maturation ovocytaire. Dans les follicules en croissance, le 4HC diminue significativement les taux de survie et de maturation; et retarde le développement folliculaire, indépendamment du traitement aux GnRHa. La chimiothérapie diminue le nombre de cellules de la granulosa par follicule tandis que la production d’adénosine monophosphate cyclique (AMPc) par million de cellules de la granulosa n'est pas modifiée, ni par le 4HC, ni par les GnRHa. La sécrétion d'oestradiol tend à être retardée dans le groupe traité à l’agoniste mais pas dans le groupe antagoniste. De même, dans les ovaires PND4, le 4HC induit une perte folliculaire importante et atteint directement les cellules de la granulosa des follicules ovariens. Aucune différence dans la distribution folliculaire, la prolifération ou l'apoptose n'a été observée entre les groupes traités avec le 4HC, peu importe la présence des GnRHa ou non. Pour conclure, en se basant sur des modèles murins robustes et originaux, notre travail remet en question l'efficacité des GnRHa pour préserver l'ovaire contre les dommages causés par la chimiothérapie que ce soit par une action directe sur l'ovaire, ou indirectement par l'absence de FSH. D'autres investigations seront nécessaires pour comprendre les mécanismes d'action potentiels des GnRHa sur l'ovaire et les voies impliquées. Des preuves expérimentales sont encore indispensables pour clore le débat sur cette option attrayante de préservation de la fertilité. / Doctorat en Sciences biomédicales et pharmaceutiques (Médecine) / info:eu-repo/semantics/nonPublished

La chimiothérapie induit une baisse de la fertilité en exerçant une toxicité directe sur les ovaires entrainant une diminution du stock ovocytaire. Couramment utilisé dans le traitement du cancer du sein, le cyclophosphamide (Cy) est une des drogues reconnue comme la plus gonadotoxique. Récemment, il a été proposé que le Cy provoquait une déplétion folliculaire par une entrée en croissance massive des follicules au repos. L’Hormone Anti-Müllérienne (AMH) étant un des facteurs régulant la sortie des follicules primordiaux de la réserve ovarienne, nous avons émis l’hypothèse que cette hormone pourrait limiter la gonadotoxicité du Cy. Nous avons montré qu’un traitement par AMH recombinante chez des souris pubères traitées par Cy, permettait effectivement de limiter la déplétion folliculaire. Sur le plan fondamental, nous avons mis en évidence que l’autophagie pouvait être un des mécanismes impliqués dans l’inhibition du recrutement folliculaire exercée par l’AMH. Enfin, la méthode de référence consistant à évaluer la réserve ovarienne chez la souris étant particulièrement longue et fastidieuse, nous avons développé une technique de comptage folliculaire automatisé en utilisant des méthodes d’intelligence artificielle, et plus particulièrement une approche de « deep learning». / Chemotherapy induces infertility by exerting a direct toxicity on the ovaries, resulting in a depletion of the follicular stockpile. Cyclophosphamide (Cy), widely used for breast cancer, is recognized as one of the most gonadotoxic agent. Recently, it has been proposed that Cy gives rise to follicular depletion by a massive growth of resting follicles which are then destroyed. Since Anti-Müllerian Hormone (AMH) is one of the factors regulating primordial follicles activation, we hypothesized that this hormone might limited Cy-induced gonadotoxicity. We have shown in pubertal mice that recombinant AMH injections are able to preserve primordial follicle loss Cy-induced and might improve fertility outcome after treatment. In addition, we provide evidence that autophagy could be one of the mechanisms involved in the inhibition of follicular recruitment by AMH. At least, nowadays, the “gold standard” method of evaluating ovarian reserve in mouse is a process particularly time-consuming and tedious. We developed a new methodology of automatic primordial follicles detection and counting within mouse ovaries, using modern artificial intelligence methods, especially deep learning approach.