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Chapter 21 The cryopreservation of preembryos
Chapter 21 The cryopreservation of preembryos BACKGROUND
Since the initiation of human IVF procedures, improving the pregnancy rates has been the goal of investigators in the field. In recent years, it has been demonstrated that the use of multiple preembryos at the time of transfer does seem to increase the clinical pregnancy rate per treatment cycle (Lopata, 1983; Jones, 1984; Marrs, 1985). The use of larger numbers of preembryos, however, incurs the increased risk of multiple pregnancy. It has been shown that the risk of multiple gestation is increasing along with the increased rate of clinical pregnancies in IVF procedures. It is currently estimated that as many as 30% of pregnancies may result in multiple gestation after multiple preembryo replacement (Society for Assisted Reproductive Technology, 1993). It is because of this increasing risk, plus the improved ability to produce preembryos, that the desire to store preembryos safely for use in subsequent cycles is of great interest.
INDICATIONS
The primary medical indication for the use of cryopreservation techniques would be for those preembryos exceeding the safe number for transfer to an individual patient. Safety should be defined as the number of preembryos that give optimal pregnancy success on a per-cycle basis without significantly increasing the risk of triplet or quadruplet gestation (chapter 11). Therefore, the number of preembryos used per transfer cycle will vary according to the pregnancy success rate per treatment cycle and the incidence of multiple gestation. The second indication, from a theoretic viewpoint, would be the possibility of increasing the success rate with IVF if preembryos could be safely cryopreserved and used for replacement when the patient was in a spontaneous ovulatory cycle. This would remove the potential risk of endometrial asynchrony, which might be present during the stimulated operative cycle. 568
Supplement 1
A third consideration would be to decrease the number of stimulated and operative oocyte recovery cycles needed for attainment of pregnancy. The use of excess or extra preembryos stored after stimulation would decrease the number of treatment cycles necessary for pregnancy initiation, because after the preembryos are thawed, a simple transfer is all that is necessary for an attempted pregnancy. Moreover, as IVF clinical pregnancy rates improve, preembryos stored from patients not desiring a second pregnancy could be used for transfer to a woman who cannot produce her own gametes (chapter 18). Finally, the ability to perform genetic screening on the preembryos for specific indications is feasible. A genetic screening to determine the normalcy of the preembryo could be done before implantation. In cases where such screening has occurred, the need for other forms of prenatal diagnosis (chorionic villus sampling or amniocentesis) and possible pregnancy termination would not be necessary. In this clinical experiment, it is essential to confirm the original diagnosis with amniocentesis or villus biopsy. With the use of preembryo genetic screening, by whatever technology available, preembryos could be frozen and stored for a short period, until the results of the genetic analyses were available. If the preembryos were found to be free of diagnosable genetic defects, they could then be thawed and transferred. The primary advantage of this method would be a decrease in the emotional and physical trauma to the patient that occurs when a genetic abnormality is identified after implantation and growth of the fetus. RESERVATIONS CONCERNING THE TECHNIQUES OF CRYOPRESERVATION
The primary reservation concerning the use of cryopreservation techniques is the risk of cryo-injury. The specific risk is currently unknown; however, from published reports, it appears that only Fertility and Sterility
50% to 60% of human preembryos may be viable after freezing and thawing (Mohr, 1985; Cohen, 1985; Marrs, 1986). Currently, cryopreservation techniques are blamed for the loss of preembryo viability. However, it may be found in the future that cryopreservation itself is not the damaging cause but that preembryo quality before freezing may not have been optimal in a significant percentage of the nonviable preembryos. This theory can be supported from human preembryo data reported in 1981. It was estimated that only 40% of preembryos initiated by IVF techniques would reach the blastocyst stage in the culture environment (Edwards, 1981). Thus, 60% of the oocytes initially fertilized in vitro would not develop to the blastocyst stage. This finding was supported by another study performed in Australia, in which only 30% to 40% of oocytes fertilized in vitro had the potential to reach blastocyst development (Lopata, 1983). If these data are accurate, then the preembryo loss seen after preembryo freezing and storage may not be due solely to the cryopreservation technique but may be related to the ultimate viability of the preembryo in an IVF system. This theory is supported by reports of improved bovine embryo viability after in vivo fertilization and freezing in comparison to freezing of in vitro embryos (Edwards, 1985). Another potential risk with cryopreservation is the unknown risk of birth defects in offspring produced by the freezing and storage of the preembryo. Currently, there are not enough live births resulting from human preembryo freezing to address this question. In the cattle industry, with its widespread use of freezing cattle blastocysts and then transferring them to recipients, birth defects have not increased. However, it must be noted that cattle embryos are fertilized in vivo and then removed from the reproductive tract for cryopreservation. A further reservation is the fear of failure in the mechanical support systems, either during the freezing process or during storage, with resultant preembryo loss. To protect against such mechanical failure, adequate backup systems for freezing and storage should be investigated. A final concern with the technique of preembryo freezing is the legal status of the preembryo. The questions of ownership and inheritance rights of preembryos have been identified but not resolved. Moreover, the highly technical procedure becomes an unnatural intrusion into the reproductive process. The concern over mechanization and manipuVol. 62, No.5, November 1994
lation of the reproductive process raises many issues from a moral standpoint, when human life forms are placed in a suspended state of deepfreeze. Furthermore, after the preembryo is frozen, short or long intervals of time may elapse before it is used. If time limitations are not considered, potential problems with family structure and societal structure may arise. RATIONALE FOR THE PROCEDURES
As previously stated, the risk and the major reservation of the freezing technique is cryo-injury to the embryo. In the human system this risk is unknown, because the viability of the preembryo in general is uncertain. Because of the lack of long-term data with human beings, it is important that good clinical investigation of the process of human preembryo cryopreservation and storage be carried out. To date, the viable births from human preembryo freezing and storage have not shown an excess rate of abnormalities. Therefore, the risk of physical defects which might be delayed in their manifestation after freezing and storage is still unknown. The question cannot be answered until more live births result from the procedure and adequate follow-up is carried out. The compelling force for using this type of procedure is to provide a more efficient system for initiating pregnancy in couples undergoing IVF. With the ability to freeze and store preembryos safely, the number of preembryos replaced per treatment cycle can be limited, and therefore the risk of multiple gestation can be limited. In addition, repetitive transfer or replacement cycles can be used without the couple being put through additional stimulation or operative manipulation for oocyte collection. As technology in genetic screening improves, preembryo freezing may be a necessary step in the assessment of the preembryo; therefore, safe cryotechniques are important. The mechanical process of human preembryo storage has to be made as safe as possible. This means that for the procedure to be performed, a backup freezing system would have to be available to ensure no interruption in the freezing process. Moreover, quality control in the laboratory during storage must be strict. Maintenance of adequate levels of liquid nitrogen in the chambers, as well as precise logging and identification of the preembryos in storage, should be provided as part of the quality control system. With these safeguards, the Supplement 1
578
risk of technical failure should be almost completely avoided and the procedure thereby justified from a mechanical viewpoint. Finally, the use of this technology for storage of a potentially viable human preembryo still has to be addressed and considered from a moral and ethical stance. Therefore, an extensive informed consent process should be undertaken with the couple, describing the risk of preembryo injury and the conditions of use after storage. Storage should be continued only as long as the normal reproductive span of the oocyte donor or only as long as the original objective of the storage procedure is in force. Therefore, transfer ofpreembryos from one generation to another would be considered unacceptable. A formal discussion with the couple should take place so that disposition of embryos not transferred to the donors can be arranged before cryopreservation. At the discretion of the donor couple, preembryos not used by the donors could therefore be transferred to infertile couples, used for approved research, examined, or discarded. If premature loss of reproductive function occurs, either by surgical procedure or ovarian failure, arrangements for extended storage may be necessary to meet the circumstances of individual situations. DISPOSITION OF CRYOPRESERVED PREEMBRYOS
With respect to decisions about the disposition of cryopreserved preembryos, the gamete providers (or those to whom they transfer their gametes) should have primary decisional authority, within the limits set by institutional policies and applicable law. The gamete providers thus may jointly decide whether their cryopreserved embryos will be thawed for transfer, thawed and not transferred, donated for implantation, donated to an institution or program for research or other uses, transferred to another location, or otherwise disposed of.* To minimize disputes and to implement the decisional authority of the gamete providers, the program or storage facility should offer them the opportunity to state, before cryopreservation is performed, dispositional directives if certain stated contingencies occur. Such directives should specify
* In consistency with my footnote in chapter 26, I believe that the decisional power of gamete providers and institutional representatives should be limited by national policy (RAM). 588
Supplement 1
the dispositional options which the gamete providers jointly authorize in case of death, divorce, unavailability, disagreement, passage of time, arrears in paying storage fees, or other stated contingencies. Options should include all dispositions that are available under applicable law and institutional policy, including the designation of a decision-maker (who may be one of the gamete providers) to exercise the decisional authority of the gamete providers. Such directives should be operative whenever a decision concerning cryopreserved preembryos must be made and the stated contingency has occurred. The directives should define, to the extent possible, the meaning of operative terms such as unavailability, disagreement, and nonpayment of storage fees. The directives should reserve the gamete providers' right jointly to alter their dispositional instructions if the stated contingencies have not occurred. Because of the importance of such dispositional directives for gamete providers and programs, the program or facility offering cryopreservation should inform or counsel couples about the various dispositional alternatives available to them and the consequences of their choice. It should give them sufficient time and information to make a considered choice about directives for future disposition ofpreembryos, including the options that the facility is free to pursue if the gamete providers have not given advance joint instructions and are unavailable or unable to agree on preembryo disposition at a later time.
COMMITTEE CONSIDERATIONS AND RECOMMENDATIONS
From a medical viewpoint, the Committee believes that cryopreservation of human preembryos is a relatively successful clinical application. The long-term risks and benefits of the procedure, although not fully assessed at present, would allow us to draw the conclusion that in a majority of cases this appears to be safe. There are advantages of preembryo cryopreservation: decreasing the need for COH, decreasing cost, decreasing the risk of multiple pregnancies, and decreasing the number of oocyte recovery cycles for a pregnancy to occur with IVF. These advantages are so compelling that the Committee believes that cryopreservation capacity is an essential component of all programs offering IVF. The rationale for this position is that the total Fertility and Sterility
reproductive potential for IVF is substantially affected by an efficacious cryopreservation program (Society for Assisted Reproductive Technology, 1993). Nevertheless, the Committee recognizes that several issues related to the disposition of unused cryo-
Vol. 62, No.5, November 1994
preserved preembryos remain morally sensitive. It is recommended that prefreeze dispositional directives be cognizant of these factors and consistent with Institutional Review Board requirements if any dispositional option is selected which can be considered experimental.
Supplement 1
598
Since the initiation of human IVF procedures, improving the pregnancy rates has been the goal of investigators in the field. In recent years, it has been demonstrated that the use of multiple preembryos at the time of transfer does seem to increase the clinical pregnancy rate per treatment cycle (Lopata, 1983; Jones, 1984; Marrs, 1985). The use of larger numbers of preembryos, however, incurs the increased risk of multiple pregnancy. It has been shown that the risk of multiple gestation is increasing along with the increased rate of clinical pregnancies in IVF procedures. It is currently estimated that as many as 30% of pregnancies may result in multiple gestation after multiple preembryo replacement (Society for Assisted Reproductive Technology, 1993). It is because of this increasing risk, plus the improved ability to produce preembryos, that the desire to store preembryos safely for use in subsequent cycles is of great interest.
INDICATIONS
The primary medical indication for the use of cryopreservation techniques would be for those preembryos exceeding the safe number for transfer to an individual patient. Safety should be defined as the number of preembryos that give optimal pregnancy success on a per-cycle basis without significantly increasing the risk of triplet or quadruplet gestation (chapter 11). Therefore, the number of preembryos used per transfer cycle will vary according to the pregnancy success rate per treatment cycle and the incidence of multiple gestation. The second indication, from a theoretic viewpoint, would be the possibility of increasing the success rate with IVF if preembryos could be safely cryopreserved and used for replacement when the patient was in a spontaneous ovulatory cycle. This would remove the potential risk of endometrial asynchrony, which might be present during the stimulated operative cycle. 568
Supplement 1
A third consideration would be to decrease the number of stimulated and operative oocyte recovery cycles needed for attainment of pregnancy. The use of excess or extra preembryos stored after stimulation would decrease the number of treatment cycles necessary for pregnancy initiation, because after the preembryos are thawed, a simple transfer is all that is necessary for an attempted pregnancy. Moreover, as IVF clinical pregnancy rates improve, preembryos stored from patients not desiring a second pregnancy could be used for transfer to a woman who cannot produce her own gametes (chapter 18). Finally, the ability to perform genetic screening on the preembryos for specific indications is feasible. A genetic screening to determine the normalcy of the preembryo could be done before implantation. In cases where such screening has occurred, the need for other forms of prenatal diagnosis (chorionic villus sampling or amniocentesis) and possible pregnancy termination would not be necessary. In this clinical experiment, it is essential to confirm the original diagnosis with amniocentesis or villus biopsy. With the use of preembryo genetic screening, by whatever technology available, preembryos could be frozen and stored for a short period, until the results of the genetic analyses were available. If the preembryos were found to be free of diagnosable genetic defects, they could then be thawed and transferred. The primary advantage of this method would be a decrease in the emotional and physical trauma to the patient that occurs when a genetic abnormality is identified after implantation and growth of the fetus. RESERVATIONS CONCERNING THE TECHNIQUES OF CRYOPRESERVATION
The primary reservation concerning the use of cryopreservation techniques is the risk of cryo-injury. The specific risk is currently unknown; however, from published reports, it appears that only Fertility and Sterility
50% to 60% of human preembryos may be viable after freezing and thawing (Mohr, 1985; Cohen, 1985; Marrs, 1986). Currently, cryopreservation techniques are blamed for the loss of preembryo viability. However, it may be found in the future that cryopreservation itself is not the damaging cause but that preembryo quality before freezing may not have been optimal in a significant percentage of the nonviable preembryos. This theory can be supported from human preembryo data reported in 1981. It was estimated that only 40% of preembryos initiated by IVF techniques would reach the blastocyst stage in the culture environment (Edwards, 1981). Thus, 60% of the oocytes initially fertilized in vitro would not develop to the blastocyst stage. This finding was supported by another study performed in Australia, in which only 30% to 40% of oocytes fertilized in vitro had the potential to reach blastocyst development (Lopata, 1983). If these data are accurate, then the preembryo loss seen after preembryo freezing and storage may not be due solely to the cryopreservation technique but may be related to the ultimate viability of the preembryo in an IVF system. This theory is supported by reports of improved bovine embryo viability after in vivo fertilization and freezing in comparison to freezing of in vitro embryos (Edwards, 1985). Another potential risk with cryopreservation is the unknown risk of birth defects in offspring produced by the freezing and storage of the preembryo. Currently, there are not enough live births resulting from human preembryo freezing to address this question. In the cattle industry, with its widespread use of freezing cattle blastocysts and then transferring them to recipients, birth defects have not increased. However, it must be noted that cattle embryos are fertilized in vivo and then removed from the reproductive tract for cryopreservation. A further reservation is the fear of failure in the mechanical support systems, either during the freezing process or during storage, with resultant preembryo loss. To protect against such mechanical failure, adequate backup systems for freezing and storage should be investigated. A final concern with the technique of preembryo freezing is the legal status of the preembryo. The questions of ownership and inheritance rights of preembryos have been identified but not resolved. Moreover, the highly technical procedure becomes an unnatural intrusion into the reproductive process. The concern over mechanization and manipuVol. 62, No.5, November 1994
lation of the reproductive process raises many issues from a moral standpoint, when human life forms are placed in a suspended state of deepfreeze. Furthermore, after the preembryo is frozen, short or long intervals of time may elapse before it is used. If time limitations are not considered, potential problems with family structure and societal structure may arise. RATIONALE FOR THE PROCEDURES
As previously stated, the risk and the major reservation of the freezing technique is cryo-injury to the embryo. In the human system this risk is unknown, because the viability of the preembryo in general is uncertain. Because of the lack of long-term data with human beings, it is important that good clinical investigation of the process of human preembryo cryopreservation and storage be carried out. To date, the viable births from human preembryo freezing and storage have not shown an excess rate of abnormalities. Therefore, the risk of physical defects which might be delayed in their manifestation after freezing and storage is still unknown. The question cannot be answered until more live births result from the procedure and adequate follow-up is carried out. The compelling force for using this type of procedure is to provide a more efficient system for initiating pregnancy in couples undergoing IVF. With the ability to freeze and store preembryos safely, the number of preembryos replaced per treatment cycle can be limited, and therefore the risk of multiple gestation can be limited. In addition, repetitive transfer or replacement cycles can be used without the couple being put through additional stimulation or operative manipulation for oocyte collection. As technology in genetic screening improves, preembryo freezing may be a necessary step in the assessment of the preembryo; therefore, safe cryotechniques are important. The mechanical process of human preembryo storage has to be made as safe as possible. This means that for the procedure to be performed, a backup freezing system would have to be available to ensure no interruption in the freezing process. Moreover, quality control in the laboratory during storage must be strict. Maintenance of adequate levels of liquid nitrogen in the chambers, as well as precise logging and identification of the preembryos in storage, should be provided as part of the quality control system. With these safeguards, the Supplement 1
578
risk of technical failure should be almost completely avoided and the procedure thereby justified from a mechanical viewpoint. Finally, the use of this technology for storage of a potentially viable human preembryo still has to be addressed and considered from a moral and ethical stance. Therefore, an extensive informed consent process should be undertaken with the couple, describing the risk of preembryo injury and the conditions of use after storage. Storage should be continued only as long as the normal reproductive span of the oocyte donor or only as long as the original objective of the storage procedure is in force. Therefore, transfer ofpreembryos from one generation to another would be considered unacceptable. A formal discussion with the couple should take place so that disposition of embryos not transferred to the donors can be arranged before cryopreservation. At the discretion of the donor couple, preembryos not used by the donors could therefore be transferred to infertile couples, used for approved research, examined, or discarded. If premature loss of reproductive function occurs, either by surgical procedure or ovarian failure, arrangements for extended storage may be necessary to meet the circumstances of individual situations. DISPOSITION OF CRYOPRESERVED PREEMBRYOS
With respect to decisions about the disposition of cryopreserved preembryos, the gamete providers (or those to whom they transfer their gametes) should have primary decisional authority, within the limits set by institutional policies and applicable law. The gamete providers thus may jointly decide whether their cryopreserved embryos will be thawed for transfer, thawed and not transferred, donated for implantation, donated to an institution or program for research or other uses, transferred to another location, or otherwise disposed of.* To minimize disputes and to implement the decisional authority of the gamete providers, the program or storage facility should offer them the opportunity to state, before cryopreservation is performed, dispositional directives if certain stated contingencies occur. Such directives should specify
* In consistency with my footnote in chapter 26, I believe that the decisional power of gamete providers and institutional representatives should be limited by national policy (RAM). 588
Supplement 1
the dispositional options which the gamete providers jointly authorize in case of death, divorce, unavailability, disagreement, passage of time, arrears in paying storage fees, or other stated contingencies. Options should include all dispositions that are available under applicable law and institutional policy, including the designation of a decision-maker (who may be one of the gamete providers) to exercise the decisional authority of the gamete providers. Such directives should be operative whenever a decision concerning cryopreserved preembryos must be made and the stated contingency has occurred. The directives should define, to the extent possible, the meaning of operative terms such as unavailability, disagreement, and nonpayment of storage fees. The directives should reserve the gamete providers' right jointly to alter their dispositional instructions if the stated contingencies have not occurred. Because of the importance of such dispositional directives for gamete providers and programs, the program or facility offering cryopreservation should inform or counsel couples about the various dispositional alternatives available to them and the consequences of their choice. It should give them sufficient time and information to make a considered choice about directives for future disposition ofpreembryos, including the options that the facility is free to pursue if the gamete providers have not given advance joint instructions and are unavailable or unable to agree on preembryo disposition at a later time.
COMMITTEE CONSIDERATIONS AND RECOMMENDATIONS
From a medical viewpoint, the Committee believes that cryopreservation of human preembryos is a relatively successful clinical application. The long-term risks and benefits of the procedure, although not fully assessed at present, would allow us to draw the conclusion that in a majority of cases this appears to be safe. There are advantages of preembryo cryopreservation: decreasing the need for COH, decreasing cost, decreasing the risk of multiple pregnancies, and decreasing the number of oocyte recovery cycles for a pregnancy to occur with IVF. These advantages are so compelling that the Committee believes that cryopreservation capacity is an essential component of all programs offering IVF. The rationale for this position is that the total Fertility and Sterility
reproductive potential for IVF is substantially affected by an efficacious cryopreservation program (Society for Assisted Reproductive Technology, 1993). Nevertheless, the Committee recognizes that several issues related to the disposition of unused cryo-
Vol. 62, No.5, November 1994
preserved preembryos remain morally sensitive. It is recommended that prefreeze dispositional directives be cognizant of these factors and consistent with Institutional Review Board requirements if any dispositional option is selected which can be considered experimental.
Supplement 1
598