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WO2025051890A1 - Dispositif intra-utérin récupérable ouvert avec moyen adhésif - Google Patents

Dispositif intra-utérin récupérable ouvert avec moyen adhésif Download PDF

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Publication number
WO2025051890A1
WO2025051890A1 PCT/EP2024/074885 EP2024074885W WO2025051890A1 WO 2025051890 A1 WO2025051890 A1 WO 2025051890A1 EP 2024074885 W EP2024074885 W EP 2024074885W WO 2025051890 A1 WO2025051890 A1 WO 2025051890A1
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open
embryos
uterine
intra
uterine device
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Pascal Mock
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Individual
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61B—DIAGNOSIS; SURGERY; IDENTIFICATION
    • A61B17/00—Surgical instruments, devices or methods
    • A61B17/42—Gynaecological or obstetrical instruments or methods
    • A61B17/425—Gynaecological or obstetrical instruments or methods for reproduction or fertilisation
    • A61B17/435—Gynaecological or obstetrical instruments or methods for reproduction or fertilisation for embryo or ova transplantation
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61F—FILTERS IMPLANTABLE INTO BLOOD VESSELS; PROSTHESES; DEVICES PROVIDING PATENCY TO, OR PREVENTING COLLAPSING OF, TUBULAR STRUCTURES OF THE BODY, e.g. STENTS; ORTHOPAEDIC, NURSING OR CONTRACEPTIVE DEVICES; FOMENTATION; TREATMENT OR PROTECTION OF EYES OR EARS; BANDAGES, DRESSINGS OR ABSORBENT PADS; FIRST-AID KITS
    • A61F6/00—Contraceptive devices; Pessaries; Applicators therefor
    • A61F6/06—Contraceptive devices; Pessaries; Applicators therefor for use by females
    • A61F6/14—Contraceptive devices; Pessaries; Applicators therefor for use by females intra-uterine type

Definitions

  • the present invention relates to an open recoverable intra-uterine device with adhesive means for fixing one or more elements selected from the group consisting of embryos, male and/or female gametes, fertilized oocytes, unfertilized eggs, or combinations thereof, used particularly for culturing gametes and/or embryos in utero.
  • floating embryo arrives in the uterine cavity from the Fallopian tube where fertilization happened, at the fourth day of its development (morula) and around 24h before the beginning of implantation. This short period of time before implantation may play a role in the decrease of expulsion risk from the uterine cavity.
  • in vitro fertilization In in vitro fertilization (IVF), however, generally, the embryo is transferred into the uterine cavity at day 2 or 3, i.e., it is present in the uterine cavity 48 to 72 hours before its presence during the natural conception. Consequently, it remains much more time in a floating state which may contribute to a higher risk to be expelled from the uterine cavity.
  • Different reasons are usually proposed to be implicated in this risk of expulsion including uterine peristalsis and contractions, low site of deposition and negative pressure generated when removing the transfer catheter.
  • Several strategies as bed rest, fibrin sealant (US 6 196 965) and mechanical closure of the cervix were proposed to avoid this problem of embryo expulsion. Unfortunately, all these strategies failed to resolve this problem.
  • EVs extracellular vesicles
  • MVs microvesicules
  • EXOs exosome
  • RNAs IncRNA, mRNA, small non-coding RNA, rRNA, miRNA
  • DNAs DNAs.
  • endometrium itself may be implicated in the preparation of embryo to implant.
  • miR-30d containing EXOs is secreted by endometrium and becomes internalized by embryonic trophoblast cells. It has been shown to induce genes involved in embryonic implantation (Vilella et al., 2015). It was also reported that mtDNAs present in MVs which were secreted by endometrial cells, seems to be involved in the embryonic ATP production. All these possible effects from the endometrium on the embryo highlights that the reproductive tract EVs cargo has an important role in reproductive events, which is missing in current in vitro media.
  • vaginal device One of them (developped by Invocell®) is a vaginal device. It consists in a box as a real container without any link and communication with the environment and containing oocytes and sperm and placed into the vagina. Only temperature, no light and presence of patient’s movement are considered to be « in vivo faculty
  • the second one is an intra-uterine device disclosed in the application WO 03/011200. It consists in a permeable silicone capsule permitting a communication between its inner and outer part.
  • the post ICSI (intra-cytoplasm sperm injection) oocytes are placed into this capsule and transitory placed into the uterus less than 2 hours later as a mini-intrauterine device (IUD) offering to early preimplantatory embryo a natural environment.
  • IUD mini-intrauterine device
  • the principle of this device is to allows the exchanges between the uterine medium and the embryo/gametes encapsulated in the device via the wall of the device made from a permeable material. However, even if several molecules contained in the uterine medium are able to cross the permeable wall, the exchange of molecules and circulation of the uterine medium is insufficient and the embryos/gametes placed in the devices are not in optimal culture conditions.
  • a permeable porous membrane is fragile and handling of this capsule, in particular when loading and unloading encapsulated elements, or when implanting the capsule in or recovering it from the uterus, could damage the wall of the capsule and possibly cause the loss of the encapsulated elements.
  • a third device was provided (described in the international application W02007074409), wherein instead to a wall made from permeable material, the device was provided with a wall having several openings with a particular size susceptible to allow molecules of the uterine medium to enter into the device but blocking the exit of embryos/gametes placed in the device.
  • the device was provided with a wall having several openings with a particular size susceptible to allow molecules of the uterine medium to enter into the device but blocking the exit of embryos/gametes placed in the device.
  • the endometrial cells and other cells necessary to the preimplantation development of the embryo whose size is bigger than the size of the openings cannot penetrate into the device and the culture conditions are thus not optimal.
  • the high viscosity of the uterine fluid may explain why exchanges between microenvironment and uterine elements are not optimal with this device.
  • the inventor of the present invention decided to continue the effort of improvement of transitory in vivo/ in utero culture for embryo/oocyte development in order to decrease the in vitro culture time before definitive transfer in the uterine cavity, since this appears to him to be the best route for improving the methods of assisted fertilization and particularly, the functionality of embryo in order to improve the process of implantation. Maintaining the principle of using the uterine cavity as an incubator and the uterine medium as a natural culture medium, the inventor found out a solution allowing to put into very close contact (as during the natural conception), the embryo/gametes with all elements (molecules and cells) of the uterine medium without any restriction.
  • This solution consists in an open recoverable intra-uterine device which comprises a support having a surface coated with adhesive means retaining the embryos/gametes and a removable cover part.
  • the removable cover part When inserted into the uterine cavity, the removable cover part is removed and the embryos/gametes fixed on the support of the device are entirely exhibited to the uterine medium as the embryos/gametes during the natural fertilization but remain solidly fixed on the surface of the support of the device.
  • the removable cover part recovers them again.
  • the present invention thus relates to open recoverable intra-uterine device (1 ) comprising:
  • cover part (4) of the device (1 ) of the invention may be removed when the device is introduced in the uterine cavity it allows the uterine fluid even with its high viscosity with all its components (molecules and cells) to be in cellular contact with the elements fixed on the support (2).
  • the main advantage of the device of the invention is the possibility to put into direct contact the elements fixed on the support (2) with the uterine fluid without releasing these elements in the uterine cavity because they are solidly fixed on the support (2). At this manner, after predetermined period of time, these elements may be recovered. They are safely recovered since the removal cover part (4) is able to recover them again when the device is retrieved from the uterine cavity, passing by the cervix and the vagina.
  • the device (1 ) of the present invention is characterized in that the elements fixed by the adhesive means are not released in the intra-uterine cavity. This is in contrast to the intrauterine devices known in the art which are notably used for transferring embryos/spermatozoids during the assisted fertilization.
  • the contact of the fixed elements with the uterine fluid which may comprise fluids coming both from the uterine cavity and from the fallopian tubes, can favor the development of the embryos. Furthermore, exchange of cells can facilitate subsequent re-implantation and nidation of the embryo in the uterus.
  • the phenomena of interaction with the uterine fluid and all its components, enhanced compared to the state of the art, can be of a kind to favor the development of the elements fixed on the support (2) of the device (1 ), and in particular the development of the embryo or the fertilization of the implanted eggs.
  • the open recoverable intra-uterine device (1 ) of the invention is able i) to improve embryo quality and invasive phenotype and euploidy level; ii) to respect this high vulnerability period of embryo development with genome reprogramming; iii) to increase involvement of future parents, mother’s uterine cavity being used as natural biological incubator; iv) to ensure close contact with endometrial cells and uterine fluid offering real cross-talk between the mother and the embryo; v) to offer a period of rescue and wellness to embryos before or after all sophisticated handlings such as ICSI, vitrification or in vitro culture, and vi) to perform complete in vivo culture of several oocytes with later intrauterine insemination or a prolonged period of in vivo culture until the hatching process which may be considered as an in vivo assisted hatching and adhesion process when limited number of embryos are cultivated in this in vivo and natural microenvironment.
  • FIG.1 shows a diagrammatic perspective view of one embodiment of the open recoverable intrauterine device (1 ) of the invention
  • FIG.2 shows a diagrammatic perspective view of one embodiment of the open recoverable intrauterine device (1 ) of the invention when the cover part (4) is partially removed;
  • FIG.3 shows a diagrammatic perspective view of wells (3);
  • FIG.4 shows immunohistochemistry (IMH) staining of adult ovarian mice tissue.
  • DAPI was used to stain nucleus and anti-ZP2 antibody coupled with a fluorescent secondary antibody (donkey anti-rat 594) was used to stain zona pellucida.
  • FIG.5 shows immunohistology image of embryo: post-incubation with primary antibody anti-ZP2 antibody and secondary fluorescent antibody donkey anti-rat 594 to stain the zona pellucida (grey). DAPI (dark grey) was used to stain the nuclei;
  • FIG.6 shows schema of the four tested conditions on mice embryos. 181 zygotes were separated into four groups and were treated with KSOM medium (#1 ), primary antibody (#2), primary antibody then DynabeadsTM (#3) or with DynabeadsTM alone (#4). Embryo development was assessed every 24h;
  • FIG.7 shows immunohistology image after addition of secondary fluorescent antibody and DAPI. Conditions without primary antibody (#1 and #4) were not stained. Conditions #2, #3 and #5 (magnets tested every day) were stained with the secondary fluorescent antibody.
  • the open recoverable intra-uterine device (1 ) shown in FIG.1 is particularly used in assisted fertilization techniques to implant and temporarily maintain male and female gametes (in vivo fertilization) and/ or embryos (preimplantation development) in a uterine cavity.
  • this intra-uterine device enables one or more elements (same or different) to be fixed on the support (2) at the start of development, for example the embryo in the start of embryo development.
  • the device is introduced into the uterine cavity for a predetermined period (from a few hours to a few days), and thereafter it is recovered to extract the embryo(s) or fertilized oocytes to monitor the development and/or in diagnosis purpose before intra-uterine implantation.
  • the open recoverable intra-uterine device (1 ) of the invention comprises a support (2) coated with adhesive means for fixing elements (embryos, male and/or female gametes, fertilized oocytes, unfertilized eggs) and a removable cover part (4).
  • the open recoverable intra-uterine device (1 ) of the invention may have any shape adapted to be inserted into a device used for intrauterine supply, for example a catheter.
  • the device (1 ) of the invention has a rectangular or ellipsoid shape.
  • the support (2) of the open recoverable intra-uterine device (1 ) has a length comprised between 5 mm and 20 mm, preferably between 10 mm and 15 mm and more preferably, the length is 15 mm.
  • the width of the support (2) is comprised between 0.5 mm and 1.5 mm, preferably between 0.7 mm and 1 mm and more preferably, the width is 1 mm.
  • the depth of the support is comprised between 1 mm and 10 mm, preferably between 4 mm and 8 mm and more preferably, the depth is 5 mm.
  • the solid support (2) comprises one or more wells (3) in its inner part, i.e. dig in the material of the support. These wells are coated with adhesive means able to fix one or more elements selected from the group consisting of embryos, male and/or female gametes, fertilized oocytes, unfertilized eggs, or combinations thereof.
  • the support comprises wells, the wells are considered as a part of the support.
  • the shape of the wells is preferably a circular shape (as the wells (3) on FIG. 1 , 2 and 3) but they can have any other shape allowing to receive the elements to be loaded in the well.
  • the wells may have a rectangular, a square, an oval or a conical shape.
  • FIG 3 is a perspective view of a circular well (3).
  • the wells (3) of the device (1 ) of the invention have preferably a diameter comprised between 0.1 mm and 0.5 mm, more preferably between 0.1 and 0.3 and even more preferably, the diameter of the wells is 0.15 mm.
  • the diameter of the wells may vary in accordance with the shape of the wells. For example, when the wells have a conical shape, the wide part of the wells which is opened on the surface of the support, has the same diameter as those for the circular walls disclosed above.
  • the solid support (2) of the device (1 ) is constituted from a biocompatible material.
  • the materials used must be tested to be non-toxic and stable in use.
  • This material may be selected from the group consisting of a polymer, a ceramic, a glass, an elastomeric material, a stainless steel, a metal material, a titanium and a titanium alloy, preferably a titanium or a titanium alloy.
  • the support (2) can be produced in a porous polymer, of the polyethersulfone (PES), polyacrylate, acry-late copolymer or polyvinylidene type.
  • the device (1 ) of the invention also comprises a removable cover part (4).
  • the cover part (4) is partially removed.
  • This removable cover part aims to protect the element fixed on the support (2) of the device or loaded in the wells (3) during the insertion of the device (1 ) in the means (for example a catheter, particularly a catheter which allows to maintain the cover part folded on the device during the insertion in the vagina and before attending the uterine cavity) adapted to be inserted into uterine cavity, wherein the device (1 ) of the invention is released.
  • the removable cover part (4) is partially removed such as the elements fixed on the support (2) remain fully exhibited to uterine medium.
  • the removable cover part (4) is replaced on the support (2) in order to protect the fixed elements when the device is recovered from the uterine cavity.
  • the use of a removable cover part (4) which may be folded on the surface of the support (2) when the device (1 ) is inserted in the supplying means and removed when the device (1 ) reaches the uterine cavity, allows the elements fixed on the support (2) and/or on the wells (3) to be exhibited to the uterine medium and all its components (molecules and cells) without any restriction. This allows to put these elements in direct contact with the uterine medium exactly at the same manner as during natural fertilization.
  • the use of removable cover part (4) allows to safely recover these elements before recovering them with the device (1 ).
  • the material used for manufacturing the removable cover part (4) may be any material which allows to be removed and then replaced without provoking damages on the elements fixed on the support.
  • this material may be a shape memory material.
  • a shape memory material refers to a material which can be deformed, generally when cold but returns to its pre-deformed ("remembered") shape, generally when heated.
  • the shape memory material used for manufacturing the removable cover part (4) of the device of the present invention is selected from shape memory alloy, shape memory polymers or shape memory composites.
  • the shape memory material is a biocompatible material.
  • the shape memory material is a shape memory biocompatible polymer.
  • the removable cover part (4) of the open recoverable intra-uterine device (1 ) has preferably the same dimensions as the support (2).
  • the cover part has a length comprised between 5 mm and 20 mm, more preferably between 10 mm and 15 mm and even more preferably, the length is 15 mm.
  • the width of the removable cover part (4) is comprised between 0.5 mm and 1.5 mm, preferably between 0.7 mm and 1 mm and more preferably, the width is 1 mm.
  • the depth of the removable cover part (4) is comprised between 1 mm and 10 mm, preferably between 4 mm and 8 mm and more preferably, the depth is 5 mm.
  • the removable cover part (4) has a rectangular shape with a length of 15 mm, a width of 1 mm and a depth of 5 mm.
  • the removable cover part (4) is partially removed when the device is inserted in the uterine cavity in order to put into direct contact the element fixed on the support (2) and the uterine medium and then, when the device is recovered, the removable cover part (4) is folded on the support (2) during the passage of the device in the cervix and the vagina.
  • a one extremity of the removal cover part (4) is attached to the well of the supplying means so as when the device (1 ) is ejected from the supplying means in the uterine cavity, the removal cover part (4) is removed from the support (2) allowing thus to exhibit the elements attached on the support (2) to the uterine medium. Then, when after predetermined period the device (1 ) is removed from the uterine cavity, the device (1 ) is replaced in the supplying means and the removable cover part (4) is replaced (folded) on the surface of the support (2) to cover the fixed elements.
  • the open intra-uterine device (1 ) of the present invention compared to other intra-uterine devices disclosed in the relating art is particular in that the protection of the embryo/gametes fixed on the support (2) of the device must be assured two times: 1 ) when the device is inserted in the uterine cavity and 2) when the device is recovered from the uterine cavity with the fixed embryos/gametes after predetermined period of time.
  • the removable cover part (4) must be designed so as to allow to exhibit the embryo/gamete in the uterine cavity and to cover them when they are introduced and recovered from the uterine cavity.
  • the device (1 ) with the fixed element has to be recovered from the uterine cavity, it is essential that the elements are fixed at the robust manner but at the same time this fixation has to be reversible since they are later replaced in a catheter allowing to introduce them in the uterine cavity for implantation.
  • the solid support is thus coated with adhesive means (or also called herein “adhesive compounds”).
  • adhesive means refers to means that allow the elements recited above (embryos/gametes etc.) to be adhered, to be attached or to be fixed on the support (2) of the device. Therefore, the terms “adhesive”, “adherent”, “attached” and “fixed” and terms derived from these ones are used interchangeably herein.
  • the adhesive means used in the open recoverable intra-uterine device of the invention are selected from any means able to ensure strong but reversible attachment of the loaded elements. Particularly, the adhesive means are selected from biochemical adhesive compounds or from mechanical, particularly mechanical-immuno-adhesive means (Novo S. et al., 2013).
  • the biochemical adhesive compounds are selected from the group consisting of compounds having the ability to bind receptor(s) specific to zona pellucida glycoproteins (ZP), cyanoacrylates, fibrine glue, biosensors mimicking the structure of grasshoppers’ feet and polysilicon barcodes for adhesion to the zona pellucida.
  • ZP zona pellucida glycoproteins
  • cyanoacrylates cyanoacrylates
  • fibrine glue cyanoacrylates
  • biosensors mimicking the structure of grasshoppers’ feet
  • polysilicon barcodes for adhesion to the zona pellucida.
  • the biochemical adhesive compounds coating the solid support (2) and/or the wells (3) are preferably selected according to their ability to bind receptor(s) specific to zona pellucida glycoproteins (ZP) that surrounds the mammalian oocytes (3D) until hatching of the blastocysts (at day 5 in the human embryo). Structural basis of egg coat-sperm recognition at fertilization may help to find the best receptor present on the sperm’s head to coat the support (2) and/or the wells (3) of the device (1 ) of the invention to increase adherence of embryos/gametes on it.
  • Compounds binding ZP on oocytes and/or embryos which may be used for coating the support and/or the wells of the device of the invention are selected from:
  • beta-1 ,4-galactosyltransferase (GalT1 ) with its ability to bind N-acetylglucosamine residues on ZP;
  • WGA wheat germ agglutinin
  • GlcNAc N-acetylglucosamine
  • sialic acid carbohydrates that are abundant on the surface of the plasma membrane and zona pellucida (3C) of oocytes and embryos.
  • biochemical adhesive means are selected from:
  • - cyanoacrylates such as N-butyl cyanoacrylate and 2-Octyl cyanoacrylate (also called Dermabond disclosed in Min et al., 2011 );
  • the adhesive means are mechanical-immuno-adhesive means.
  • the mechanical means is a support or a stand and biochemical means is an antibody capable of binding at least one protein of zone pellucid of embryos, female gametes, fertilized oocytes or unfertilized eggs.
  • the support or strand is a bead and the mechanical- immuno-adhesive means is a bead coated with said antibody.
  • the term “support” or “stand” is used to design a mechanical means which is a part of the adhesive means.
  • the “support” is not the same as the support (2) which is included in the structure of the device (1 ) of the invention and on which surface are fixed the adhesive means.
  • the support or stand, particularly the bead may be manufactured from any material provided that it is able to be coated with an antibody anti zone pellucid.
  • a material is a material having an attractive force.
  • the bead coated with an antibody anti zone pellucid is a magnetic bead which is retained on the solid support (2) and/or the surface of the wells (3) by a means providing a magnetic force.
  • the magnetic beads are fixed on the surface of the solid support (2) and/or on the surface of the wells (3) of the device via a metal element providing a magnetic force.
  • a means creating a magnetic field may be used to fix the magnetic beads on the solid support of the device (1 ).
  • the support (2) and/or wells (3) are coated with magnetic beads coated with antibody anti-zone pellucid selected from the group consisting in anti-ZP-1 , anti- ZP-2 and anti-ZP-3, particularly ZP-2.
  • the inventor developed a protocol to bind antibodies (primary) to the zona pellucida of embryo in a non-toxic manner.
  • the binding of magnetic beads for example DynabeadsTM
  • the inventor surprisingly found that coupling directly the anti-ZP2 antibodies to the magnetic beads and maintain a continuous motion, assures the binding of the magnetic beads throughout the zona pellucida in a more homogenous and efficient manner, which results in a higher magnetic retrieval yield.
  • the adhesive means is magnetic bead and the anti-ZP antibody is anti-ZP-2 antibody.
  • the biding is performed under gently stirring allowing to create a motion of the beads.
  • the adhesive means are mechanical means such as 3D printed structures able to retain the above recited elements, particularly during their exhibition in the uterine cavity.
  • a particular mechanical means is for example a cage wherein the openings formed by the bars crossing have a size smaller than the size of the element putted into the cage. Therefore, the element, when loaded in the cage cannot be released in the uterine cavity but the uterine fluid with all substances necessary to the in-utero culture can penetrate the cage and thus being into direct contact with the loaded element.
  • This cage is provided with a removable door allowing to load the element before introducing the device (1 ) in the uterine cavity and to remove the element from the cage after retrieving the device (1 ) from the uterine cavity.
  • the cage may be manufactured from a nontoxic biocompatible material. This material may be selected from the group consisting of a polymer, a ceramic, a glass, an elastomeric material, a stainless steel, a metal material, a titanium and a titanium alloy. According to one embodiment, the cage maybe manufactured from shape memory material allowing thus to load/remove the elements by the openings formed by the crossing bars. In this case, the presence of a door is not necessary.
  • the cage may have any shape adapted to the inserted element.
  • the cage may have a square shape, a rectangle shape, a circular shape, an oval shape or conical shape.
  • the cage is fixed on the support (2) of the device (1 ) or when present, in the wells (3) of the device (1 ) by any means allowing a strong fixation.
  • the cage may be extruded from the material of the support (2).
  • the elements attached on the surface of the solid support (2) and/or the wells (3) of the device of the invention are selected from the group consisting of embryos, male and/or female gametes, fertilized oocytes, unfertilized eggs, or combinations thereof.
  • these elements are obtained from mammal selected from the group consisting of bovine, ovine, porcine, horses and human. More particularly, the mammal is a human being.
  • embryo refers to mammalian fertilized oocyte (zygote), wherein the first mitosis has not been yet performed.
  • the male and female gametes are haploid reproductive male and female cells respectively.
  • the term “fertilized oocyte” refers to mammalian female gamete fertilized by a mammalian male gamete, i.e. the fertilized oocyte corresponds to a zygote (diploid cell).
  • the fertilized oocyte may be transferred via the device (1 ) of the invention into the uterine cavity for a period of 24 hours or 2 to 5 days (for attending embryo stage of development) and then, it may be retrieved to be analyzed. After such an analysis, if appropriate, the formed embryo is transferred via an appropriate device (catheter) into the uterine cavity in order to proceed to its implantation in the uterine wall.
  • the present invention also relates to a method of preparing an open retrievable intra-uterine device for placing one or more elements selected from the group consisting of embryos, male and/or female gametes, fertilized oocytes, unfertilized eggs, or combinations thereof comprising the steps of: providing said element under the appropriate form to be attached on the surface of the solid support (2) and/or the wells (3) of the device, providing the open retrievable intra-uterine device (1 ) suitable for receiving said element(s) according to the invention, attached said element(s) on the surface of the solid support (2) and/or the wells (3) of the device (1 ).
  • the open recoverable intra-uterine device of the invention furthermore cooperates with accessories (means) for placing the device in the uterine cavity.
  • accessories for placing the device in the uterine cavity.
  • a transfer catheter having an inside diameter of 1 cm to 1.3 cm may be used.
  • a catheter of this kind is adapted to pass through the cervical canal, which generally has a diameter of about 1 .5 cm.
  • the device of the present invention may also use as implantation site in the Fallopian tube.
  • This implantation needs to use a surgical procedure as coelioscopy with general anaesthesia or culdoscopy with local anaesthesia.
  • Such in vivo intra Fallopian embryo culture is similar to GIFT or ZIFT except the fact that using of the open recoverable intra-uterine device of the invention, the incubation time is under control and an unlimited number of embryos can be loaded which were retrieved and selected for transfer after a simple flushing procedure.
  • the device of the invention presents a number of advantages: said device is not likely to cause any trouble to the uterus (little or no tissue reaction, without inducing inflammatory or fibrotic reactions and or inappropriate uterine wall tissue damage or scarring) due to the fact that it is not implanted within the uterine wall, it does not require absolutely surgery or anaesthesia to be inserted and it can be inserted in a completely ambulatory fashion.
  • the device of the invention is designed such that it has a means of remaining within the uterus (i.e. small suture thread glued into the tip of the device for attachment inside or outside the uterus) and that can be easily retrieved (attached suture thread) at any time after uterine implantation.
  • the open recoverable intra-uterine device of the invention is able to present gametes and/or embryos to the natural microenvironment of the uterine cavity without encapsulation (as the similar devices in the relating art). It permits to obtain for the gametes and the embryos an optimal micro-environment in a small volume.
  • the device of the invention offers to developing embryos a closer biophysics-chemical environment with less osmolarity alteration and permits to the uterus to become a real transitory natural incubator offering the possibility of a complete physico-chemical and cellular complex cross talk to take place between the mother and embryos. Even, complete in vivo/in utero fertilization may take place into the uterine cavity with the performance of intrauterine insemination after the intrauterine transfer of this device fixing oocytes.
  • the device of the present invention resolves the problem of embryo(s) passage from the uterus through the inner cervical canal and their expulsion into the vagina.
  • the open recoverable intra-uterine device may be used successfully in assisted reproduction.
  • the present invention thus relates to a method of assisted reproduction, wherein the method comprises the following steps:
  • the “predetermined period” corresponds to the period or time necessary to postfertilization embryo development (conventionally performed in in vitro culture) before the definitive transfer of the embryo into uterine cavity.
  • the “predetermined period” may also correspond to the time necessary for performing in utero fertilization (i.e. fertilization in the device of the invention) when oocytes and spermatozoids are contained in the device.
  • the device may be placed in the uterine cavity by the means described above.
  • the device of the present invention may be used in in vivo fertilization by injecting prepared sperm and retrieval oocytes in the device for implanting it into the uterus. After a defined and controlled incubation time (2 hours for example) of in vivo and in utero culture, the sperm and oocytes are retrieved and zygotes and/or unfertilized oocytes are collected after a simple flushing procedure. Then, selection of zygotes for cryopreservation or in vitro culture of the remaining embryos to be transferred at day 3 is performed.
  • the device of the present invention may be also used in in vivo preimplantatory embryo development by injecting several embryos at different stage of development (i.e 6-8 cells) in the device followed by the implantation of the device into the uterine cavity during a controlled time (i.e 48 hours). After removal of the device from the uterus, the embryos at the blastocyst stage are flushed from the device and transferred into the uterine cavity using a conventional transfer catheter or delayed in a further cycle after freezing.
  • the device of the invention may be used in in vivo embryo assisted hatching.
  • Blocking buffer (BB) composed of PBS-BSA 3% diluted in Tween 0.1%
  • the Ab mixtures was always kept on ice.
  • the anti-ZP2 antibody binds to the zona pellucida a)
  • Anti-ZP-2 antibody was used as primary antibody and donkey anti-rat 594 (fluorescent dye) as secondary antibody to validate the binding of the primary antibody to the zona pellucida (FIG.4).
  • the fluorescent staining of the zona pellucida led to the conclusion that anti-ZP-2 specifically binds to ZP-2 receptor.
  • the primary antibody was thus validated (van Oss. et al. 1986).
  • the same protocol was adapted on fixed mouse embryos in suspension using Eppendorf tubes. Centrifugation was done in between staining and washing steps.
  • the protocol was done in technical duplicates. As shown on the FIG. 5, at the end of the experiment 90% of embryos from the control group were present and 93.3% from the tested embryos were present and expressed stained zona pellucida as expected. The protocol on embryos was thus validated. a) First, beads alone were tested under a microscope. Post incubation of the beads, a magnet was placed next to the slide to attract the beads. The beads attraction to magnetic fields was observed. b) Then, embryos were incubated with anti-ZP-2 and DynabeadsTM. Post-incubation, a magnet was placed next to the Petri dish : a weak attraction was observed. c) Finally, the inventor added magnetic beads directly on the embryos and added motion to the mixture. Post-incubation, a magnet was placed next to the Petri dish : an attraction was observed over time.
  • mice embryos at the single-cell stage were obtained by IVF, collected and separated into 4 experimental groups: 45 in the control group (#1 ) without any antibodies; 45 embryos incubated only with the primary antibody (1 °Ab) (#2); 46 embryos with both the 1 ° Ab and the Dynabeads (2°Ab) (#3); and 45 embryos for the last condition (#4) with only the 2°Ab.
  • an additional group (#5) was done with the 28 embryos at the two-cell stage. #5 had the same condition as #3 but was used as a test group to use the magnet on the embryos following the experiment.
  • condition #3 was done twice (once with single-cell embryos (#3) and once with 2-cells embryos (not shown)).
  • the experimental protocol is show on FIG. 6.
  • Groups #2, #3, #5 received the anti-ZP-2 antibody diluted in KSOM and were incubated 1 h at 37°C. Post incubation, the embryos were rinsed in KSOM drops twice and placed on a new Petri dish. Groups #3, #4 and #5 then received the beads diluted in KSOM and were incubated for 30min at 37° C. Post incubation, the embryos were rinsed twice and placed in new Petri dishes. Pictures were taken. All groups were placed overnight at 37° C. Condition #5 was used to test the magnet each day. Embryos were then placed at 37° C overnight as well.
  • Table 1 Embryo development assessment over time
  • condition #2, #3 and #5 showed fluorescence. This suggests that DynabeadsTM probably detached from the primary antibody and migrate to the bottom of the dish, not properly bound to the primary antibody, allowing for the fluorescent secondary antibodies to bind correctly.
  • the attachment of primary antibodies to DynabeadsTM is improved by inducing a continuous motion.

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Abstract

La présente invention concerne un dispositif intra-utérin récupérable ouvert comprenant des moyens adhésifs et utilisé notamment pour la culture de gamètes et/ou d'embryons in utero.
PCT/EP2024/074885 2023-09-05 2024-09-05 Dispositif intra-utérin récupérable ouvert avec moyen adhésif Pending WO2025051890A1 (fr)

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Citations (4)

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WO2003011200A2 (fr) 2001-08-01 2003-02-13 Pascal Mock Dispositif intra-uterin, procede de fabrication d'un tel dispositif et procede de pose d'elements actifs a l'interieur de la cavite uterine
WO2007074409A2 (fr) 2005-12-26 2007-07-05 Anecova Sa Dispositif intra-uterin recuperable
US20120095286A1 (en) * 2010-10-18 2012-04-19 Cezary Grygoruk Catheter for an embryo transfer and a carrier for a cell transport

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