P.I. Responsible: Antonio Pellicer Martínez, MD, PhD | Raúl Gomez, PhD
Phd Student: Hortensia Ferrero
OVARIAN REJUVENATION AND IMPROVEMENT OF THE OVARIAN RESERVE.
Nowadays up to 15% of European couples could be considered as infertile, and thus require assisted Reproductive Technologies (ART) to achieve their desired motherhood. European women have delayed the age to childbearing due to the socio-economic changes experienced by our society and therefore patient’s age is the main determinant of infertility, since it is well known that both quantity and quality of the oocytes from aged patients are seriously impaired. They are usually known as Poor Responders (PR), representing 9-24% of the patients attending an ART clinic and oocyte donation is the only practical option. Consequently, interventions to restore already aged or damaged gonads in PR and aged patients to enhance their reproductive potential should be developed. We aim to activate primordial dormant follicles already present in ovary from PR as well as to rejuvenate the ovarian niche to allow follicular growth by the following interventions.
- Stem Cell Ovarian auto-Transplantation (SCOT): We aim to induce ovarian rejuvenation by SCOT in women whose ovarian endowment has been partially consumed (PR). Previous studies suggest positive effects produced by BM transplant (BMT in damaged ovaries and raising the possibility that dormant follicles, already present or somatic cells within the ovarian niche, may benefit from a positive influence of the BM derived cells or soluble growth factors This concept is reinforced by the report of women who unexpectedly restore fertility after BMT.
- Ovarian Fragmentation for Follicular Activation (OFFA) in PR patients: Although the underlying mechanisms remain unknown, it seems that tissue sectioning plays a crucial role enhancing follicle development. Tissue fragmentation is an effective and easy way to inhibit the Hippo pathway: it changes intracellular tension, resulting in actin polymerization. This fact disrupts the hippo pathway and improves cell growth and survival, because increases gene expression of intracellular proteins associated to extracellular matrix signaling and apoptosis inhibitors, ending up in follicle activation and generation of mature oocytes. A clinical trial will be developed in PR to evaluate if OFFA improves the number of antral follicles.
Figure 2: Schematic representation of the scientific approaches employed in our proposal to achiever Ovarian Rejuvenation.
DEVELOPMENT OF BASIC METHODS TO IMPROVE SAFETY IN WOMEN’S FERTILITY PRESERVATION.
Survival after cancer in childhood and adolescence has increased to >85%. Thus, secondary effects of cancer treatment, such as the need for Fertility Preservation (FP) are an increasing request especially in women. The most frequent malignancies are Breast Cancer (BC), Hodgkin's Disease and Acute Leukemias (AL). Several methods of FP are currently employed, but to date cryopreservation of ovarian cortex and subsequent OrthoTransplantation (OT) when cancer has been cured, is the only method that has proved to be successful in terms of pregnancies and birth of healthy children. A major concern in OT is the contamination of ovarian tissue with cancer cells when the tissue is frozen with the subsequent reintroduction of malignant cells at OT. This project is organized in several Work Packages (WP) that deal with improvement of diagnostic methods in cryopreserved tissue, and new treatment methods to avoid reimplantation of cancer.
WP1-Detection of occult metastatic disease in ovarian tissue from BC patients: In order to evaluate the safety of ovarian tissue cryopreservation and transplant for BC, a mestastatic detection panel based on molecular methods capable to detect ocult metastasis by Real time PCR with 90% sensitivity and 100% specificity has been designed. Ovarian tissue from BC undergoing FP will be analyzed with this diagnostic tool to ensure absence of malignant cells.
WP2- Detection of occult metastatic cells from AL. The experimental design from WP1 will be employed with specific molecular markers for AL to detect contamination in the ovarian cortex obtained from young AL patients and follicular cells (granulosa and oocyte) from AL adults.
WP-3 New FP alternatives for patients with hematologic cancer. Dexamethasone has been proposed as anti-leukemic agent to treat ovarian cortex from AL young patients. Malignant cell contamination has been evaluated after treatment and xenograft into an immunodeficient animal model. After this preliminary study we will asses the minimum amount of cells capable to develop malignancies as well as the suitability of the currently used animal models.
WP4-In vitro activation (IVA), growth and maturation of primordial follicles from oncologic patients. a new method to be employed in any malignancy, including BC and AL, will be developed. It consists in activation of dormant primordial follicles from ovarian cortical tissue by incubation with PTEN inhibitors. Then IVA follicles will be growth in three different conditions: follicular culture in supplemented media, alginate gels supplemented with adipose tissue stem cells, and scaffolds of menopausal ovaries after matrix decellularization, also supplemented with stem cells. Follicular growth by morphology, follicular function by means of steroids and other proteins produced, and oocyte competence by spindle formation and molecular markers of oocyte maturation in humans, will be the endpoints to find the optimal method to obtain fertilizable disease-free oocytes in FP programs.
WP5- Ovarian stimulation strategies in patients with BC. We aim to evaluate the effects induced by Controlled Ovarian Stimulation could have on the molecular and histological profile of the tumor. Clinical follow up will be also evaluated.
- Ayudas económicas para el Desarrollo de Estudios de Investigación Clínica La Fe. Follicular activation by ovarian fragmentation in poor responder patients. 2014-2016
- PROMETEO 2014 – FASE II Ayudas para la realización de proyectos de I+D para grupos de investigación de excelencia. Nuevos métodos seguros de preservación de la fertilidad femenina en pacientes oncológicas. PROMETEO 2014/0045, by the Regional Valencian Ministry of Education.
- Mejora de la seguridad y nuevas estrategias en Preservación de la Fertilidad en pacientes oncológicas. Proyecto de de Investigación en Salud by the Spanish ministry of Economy and Competitiveness. PI13/02353.
- Optimizing ovarian tissue quality before cryopreservation: evaluation of three medulla separation methods on stromal and follicular viability, by the Dexeus Women's Health Foundation Grant 2013.
- Detection of occult metastatic disease in ovarian tissue of women with breast cancer: development of more specific methods to confirm previous findings. PROYECTOS DE I+D PARA GRUPOS DE INVESTIGACIÓN EMERGENTES GV/2014/115 by the Regional Valencian Ministry of Education.
- Sara Borrell postdoctoral grant. ISCIII-Spanish ministry of Economy and Competitiveness. CD11/0292.
- Development of new methods to improve security in Fertility Preservation. Funded by MICCIN- Spanish Government . SAF2011-30031-C02-01.
- Ayudas Complementarias para Proyectos de I+D by the Regional Valencian Ministry of Education. ACOMP/2013.
- Ayudas Complementarias para Proyectos de I+D by the Regional Valencian Ministry of Education. ACOMP/2012
- Fertility preservation for age-related fertility decline. Donnez J, Dolmans MM, Pellicer A, Diaz-Garcia C, Ernst E, Macklon KT, Andersen CY. Lancet. 2015 Feb 7;385(9967):506-7. doi: 10.1016/S0140-6736(15)60198-2.
- Ovary transplantation: to activate or not to activate. Kawamura K, Cheng Y, Sun YP, Zhai J, Diaz-Garcia C, Simon C, Pellicer A, Hsueh AJ. Hum Reprod. 2015 Nov;30(11):2457-60. doi: 10.1093/humrep/dev211. Epub 2015 Sep 6
- New methods to improve the safety assessment of cryopreserved ovarian tissue for fertility preservation in breast cancer patients. Rodríguez-Iglesias B, Novella-Maestre E, Herraiz S, Díaz-García C, Pellicer N, Pellicer A. Fertil Steril. 2015 Sep 30. pii: S0015-0282(15)01763-X. doi: 10.1016/j.fertnstert.2015.08.009. [Epub ahead of print]
- Short-Term PTEN Inhibition Improves In Vitro Activation of Primordial Follicles, Preserves Follicular Viability, and Restores AMH Levels in Cryopreserved Ovarian Tissue From Cancer Patients. Novella-Maestre E*, Herraiz S*, Rodríguez-Iglesias B, Díaz-García C, Pellicer A. PLoS One. 2015 May 29;10(5):e0127786. doi: 10.1371/journal.pone.0127786. eCollection 2015.
- Single site laparoscopy for fertility preservation: a cohort study. Núñez Valera MJ, Padilla Iserte P, Higueras García G, Herraiz S, Rubio JM, Romeu Villarroya M, Pellicer A, Díaz-García C. J Minim Invasive Gynecol. 2015 Feb;22(2):291-6. doi: 10.1016/j.jmig.2014.10.018. Epub 2014 Oct 29.
- The artificial ovary: any new step is a step forward. Díaz-García C, Herraiz S. Fertil Steril. 2014 Apr;101(4):940. doi: 10.1016/j.fertnstert.2014.01.057. Epub 2014 Mar 6.
- Improving ovarian tissue cryopreservation for oncologic patients: slow freezing versus vitrification, effect of different procedures and devices. Herraiz S, Novella-Maestre E, Rodríguez B, Díaz C, Sánchez-Serrano M, Mirabet V, Pellicer A. Fertil Steril. 2014 Mar;101(3):775-84. doi: 10.1016/j.fertnstert.2013.11.016. Epub 2013 Dec 17.
- Restoration of ovarian activity and pregnancy after transplantation of cryopreserved ovarian tissue: a review of 60 cases of reimplantation. Donnez J, Dolmans MM, Pellicer A, Diaz-Garcia C, Sanchez Serrano M, Schmidt KT, Ernst E, Luyckx V, Andersen CY. Fertil Steril. 2013 May;99(6):1503-13. doi: 10.1016/j.fertnstert.2013.03.03
Works submitted to international conferences
- American Society Reproductive Medicine (ASRM) 2015 Annual Meeting. Baltimore, Oct 2015. Díaz-García C., Herraiz S., Such E., Andrés M., Mayordomo E., Sanz J. and Pellicer A. Evaluation of anti-leukemic in vitro treatment with dexamethasone of ovarian cortex prior to transplantation
- 62nd Annual Meeting of the Society for Reproductive Investigation. San Francisco, USA, March 2015. Herraiz S; Díaz-García C; Novella-Maestre E; Rodriguez-Iglesias B; Monterde M; Pellicer A. Genomic, Proteomic and Phosphorylation Profiles of PTEN-Akt Pathway in Previously Cryopreserved Ovarian Cortex from Cancer Patients Undergoing Follicular In Vitro Activation.
- Freezing of oocytes, embryos and ovarian tissue: focus on fertility management and fertility preservation. Brussels, Belgium. Jan 2015. Herraiz S; Díaz-García C; Novella-Maestre E; Rodriguez-Iglesias B, Pellicer A. Genomic expression and phosphorylation profiles of PI3K-Akt pathway in ovarian cortex from cancer patients undergoing follicular in vitro activation with a PTEN inhibitor. Oral communication
- Freezing of oocytes, embryos and ovarian tissue: focus on fertility management and fertility preservation. Brussels, Belgium. Jan 2015 . Díaz-García Herraiz S; Such E; Andrés M; Barragán E; Martin I; Novella-Maestre E; Sanz J; Pellicer A. Evaluation of anti-leukemic in vitro treatment with Dexamethasone of ovarian cortex prior to transplantation. Oral communication
- Freezing of oocytes, embryos and ovarian tissue: focus on fertility management and fertility preservation. Brussels, Belgium. Jan 2015. Susana Monzó; Sonia Herraiz; Sonia Lozano; Antonio Pellicer; César Díaz-García. Optimizing ovarian tissue quality before cryopreservation: evaluation of three medulla separation methods on stromal and follicular viability. Oral communication.
- 30th Annual Meeting of the European Society of Human Reproduction (ESHRE). Germany, 29 Jun-2 July 2014. E. Novella-Maestre, B. Rodriguez-Iglesias, S. Herraiz, A. Verdeguer, A. Pellicer, M.M. Andrés Gonadal effects of acute leukaemia treatment in prepubertal girls. O235 ORAL PRESENTATION
- 30th Annual Meeting of the European Society of Human Reproduction (ESHRE). Germany, 29 Jun-2 July 2014. S. Lozano, S. Herráiz, S. Monzó, A. Pellicer, C. Díaz-García Optimizing ovarian tissue quality prior to cryopreservation for fertility preservation - effects of mechanical separation on cell damage.
- 3rd International Society Fertility Preservation (ISPF) World Congress. Herraiz S. Vitrification: New insights
- 29th Annual Meeting of the European Society of Human Reproduction (ESHRE). London-UK, 7-10 July 2013. Herraiz, S; Novella-Maestre,E; Rodriguez-Iglesias,B; Díaz-García, C; Mirabet, V and Pellicer, A. Cryopreservation of primordial follicles from human ovarian tissue after slow-rate freezing or vitrification and xenografting.
- 29th Annual Meeting of the European Society of Human Reproduction (ESHRE). London-UK, 7-10 July 2013. Novella-Maestre, E.; Herraiz, S.; Rodriguez-Iglesias, B; Díaz, C. and Pellicer, A. Activation of primordial follicles in vitro preserves the follicular viability and restores the AMH levels in cryopreserved ovarian tissue. Poster.
- 29th Annual Meeting of the European Society of Human Reproduction (ESHRE). London-UK, 7-10 July 2013. Rodriguez-Iglesias, B.; Herraiz, S.; Novella-Maestre, E.; Díaz, C.; Vera, F. and Pellicer, A. Safety assessment of ovarian cryopreservation by molecular techniques in Breast Cancer Patients
- 60th Annual Scientific Meeting of the Society of Gynaecologic Investigation. Orlando, USA. Mar 2013. Herraiz S, Rodriguez B, Díaz C, Santaballa A, Vera F Rosello E , Sánchez-Serrano M, Pellicer A. New Molecular Markers To Detect Occult Metastases In Ovarian Cortex From Breast Cancer Patients.
- 28th Annual Meeting of the European Society of Human Reproduction (ESHRE). - Istanbul, Turkey, 1-4 July 2012. S. Herraiz1, E. Novella-Maestre2, A. Pellicer3. Improving ovarian tissue cryopreservation: slow freezing vs. vitrification, effect of different procedures and devices. ORAL PRESENTATION
- 28th Annual Meeting of the European Society of Human Reproduction (ESHRE). Istanbul, Turkey, 1-4 July 2012. E. Novella-Maestre1, S. Herraiz2, J.M. Vila3, A. Pellicer3. Effect of antiangiogenic treatment on peritoneal endometriosis-associated nerve fibers. ORAL PRESENTATION
- 59th Annual Scientific Meeting of the Society of Gynaecologic Investigation. Mar 2012, San Diego, USA. Sonia Herraiz, Patricia Blanco, César Díaz, Patrocinio Polo, José M Rubio, Antonio Pellicer, Edurne Novella-Maestre. In vitro activation of cryopreserved ovarian cortex follicles from oncologic patients. ORAL PRESENTATION