Successful haematopoietic stem cell transplantation in 44 children from healthy siblings conceived after preimplantation HLA matching.

Haematopoietic stem cell transplantation (HSCT) remains the best therapeutic option for many acquired and inherited paediatric haematological disorders. Unfortunately, the probability of finding an HLA matched donor is limited. An alternative technique is PGD combined with HLA matching, which offers the possibility of selecting unaffected embryos that are HLA compatible with the sick child, with the aim of possible use of stem cells from the resulting baby in future. Since the first successful report for Fanconi anaemia a decade ago, the therapeutic success of this technique was reported in a few cases and for a limited number of disorders. Here, we report full recovery of 44 sick children who received HSCT from healthy infants conceived after pre-implantation HLA matching for the following 10 indications; beta-thalassaemia, Wiskott-Aldrich syndrome, Fanconi anaemia, sickle cell anaemia, acute myeloid leukaemia, acute lymphoblastic leukaemia, Glanzmann's thrombasthaenia, Diamond-Blackfan anaemia, X-linked adrenoleukodystrophy and mucopolysaccharidosis type I. No serious complications were observed among recipients and donors. Graft failure occurred in four children with beta-thalassaemia where a second HSCT was planned. Preimplantation HLA matching is a reliable technique and provides a realistic option for couples seeking treatment for an affected child when no HLA-matched donor is available.

Seven years of experience of preimplantation HLA typing: a clinical overview of 327 cycles.

Preimplantation human leukocyte antigen (HLA) typing allows the birth of healthy children who are potential donors of stem cells for their affected siblings. This technique can be used for acquired diseases such as leukaemia or can be used for single-gene disorders such as thalassaemia. This retrospective study presents clinical data obtained from 171 couples who had undergone 327 preimplantation HLA typing cycles: 262 cycles for HLA typing in combination with mutation analysis and 65 cycles for the sole purpose of HLA typing. Of the diagnosed embryos 17.6% were found to be HLA matched. Embryo transfer was performed in 212 cycles, 34.9% clinical pregnancy rate per transfer was achieved and 59 healthy and HLA-compatible children were born. Twenty-one sick children have been cured through haemopoietic stem cell transplantation. The effect of maternal age and ovarian reserve on reproductive outcome was assessed retrospectively. The data demonstrated that, once a mutation-free and HLA-compatible embryo was found, clinical pregnancy rate did not differ statistically significantly despite the presence of some cycle-related limitations such as advanced maternal age and/or diminished ovarian reserve. Preimplantation HLA typing is an effective therapeutic tool for curing an affected sibling even for poor-prognosis patients. Preimplantation human leukocyte antigent (HLA) typing allows the birth of healthy children who are potential donors of stem cells for their affected siblings. This technique can be used for acquired diseases such as leukaemia or can be used for single-gene disorders such as thalassaemia. This study presents clinical data obtained from 171 couples who underwent 327 preimplantation HLA-typing cycles. Of these, 262 cycles were performed for HLA typing in combination with mutation analysis and 65 cycles were performed for the sole purpose of HLA typing. A total of 17.6% of the diagnosed embryos were found to be HLA matched. Embryo transfer was performed in 212 cycles. The clinical pregnancy rate per transfer was 34.9% and 59 healthy and HLA compatible children were born. Twenty-one sick children have been cured through haemopoietic stem cell transplantation. The effect of maternal age and ovarian reserve on reproductive outcome was assessed retrospectively. The data demonstrated that, once a mutation-free and HLA-compatible embryo was found, clinical pregnancy rates did not differ statistically significantly by the presence of some cycle-related limitations such as advanced maternal age and/or diminished ovarian reserve. Preimplantation HLA typing is an effective therapeutic tool for curing an affected sibling even for poor-prognosis patients.

A genetic survey of 1935 Turkish men with severe male factor infertility.

Male factor infertility is the sole reason in approximately 25% of couples who suffer from infertility. Genetic factors such as numerical and structural chromosomal abnormalities and microdeletions of the Y chromosome might be the cause of poor semen parameters. The results of karyotype analyses and Y-chromosome microdeletions of 1935 patients with severe male factor infertility, which is the largest series from Turkey, were assessed retrospectively. The frequency of cytogenetic abnormalities among 1214 patients with non-obstructive azoospermia (NOA) and 721 patients with severe oligoasthenoteratozoospermia (OAT) were 16.40 and 5.83% respectively. The overall incidence of Y-chromosome microdeletion was 7.70%. The incidence of Y chromosome microdeletion in patients with NOA and OAT was 9.51 and 1.86% respectively. The abnormality rate increased with the severity of infertility. Some patients (n = 22) were carriers of both chromosomal abnormalities and Y-chromosome microdeletions. Results suggest the need for genetic screening and proper genetic counselling before initiation of assisted reproduction treatment.

Genetic diagnosis in infertile men with numerical and constitutional sperm abnormalities.

Infertile men having numerical or structural sperm defects may carry several genetic abnormalities (karyotype abnormalities, Y chromosome microdeletions, cystic fibrosis transmembrane conductance regulator (CFTR) gene mutations, androgen receptor gene mutations, and abnormalities seen in sperm cells) leading to this situation. First we aimed to investigate the relationship between the numerical and constitutional (morphological) sperm anomalies and the genetic disorders that can be seen in infertile males. Our other aim was to compare two different kinds of kits that we use for the detection of Y chromosome microdeletions. Sixty-three infertile males [44 nonobstructive azoospermic, 8 severe oligozoospermic, and 11 oligoasthenoteratozoospermic] were investigated in terms of somatic chromosomal constitutions and microdeletions of the Y chromosome. Sperm aneuploidy levels were analyzed by fluorescence in situ hybridization (FISH) in sperm cells obtained from the semen of six OAT patients. Microdeletion and sex chromosome aneuploidy (47,XXY) rates in somatic cells were found to be approximately 3.2% and 4.7%, respectively. Sperm aneuploidy rates were determined as 9%, 22%, and 47% in three patients out of six. Two of these three patients also had high rates of head anomalies in semen samples. High correlation was found between sperm aneuploidy rates and sperm head anomalies. Since the introduction of the assisted reproductive techniques for the treatment of severe male infertility, genetic tests and genetic counseling became very important due to the transmission of genetic abnormalities to the next generation. Thus in a very near future, for a comprehensive male infertility panel, it will be essential to include additional genetic tests, such as CFTR gene mutations, sperm mitochondrial DNA mutations, and androgen receptor gene mutations, besides the conventional chromosomal analyses, Y chromosome microdeletion detection, and sperm-FISH analyses.

Birth of a healthy infant after preimplantation genetic diagnosis by sequential blastomere and trophectoderm biopsy for ß-thalassemia and HLA genotyping.

BACKGROUND: Preimplantation genetic diagnosis (PGD) is a widely used technique for couples at genetic risk and involves the diagnosis and transfer of unaffected embryos generated through in vitro fertilization (IVF) techniques. STUDY DESIGN: For those couples who are at risk of transmitting a genetic disease to their offspring, preimplantation embryos can be selected according to their genetic status as well as human leukocyte antigen (HLA) compatibility with the affected child. Stem cells from the resulting baby's umbilical cord blood can be used for transplantation to the affected sibling without graft rejection. RESULTS: Here we report successful hematopoietic stem cell transplantation (HSCT) after the birth of a healthy infant, who was born after successful PGD testing with both cleavage stage and blastocyst stage biopsy for the purpose of diagnosis of ß-thalassemia and HLA compatibility. CONCLUSION: The specific feature of this work is not only to have the first successful HSCT achieved in Bulgaria after using preimplantation HLA typing technique, it also demonstrates how to accomplish this success via cross-border collaboration of different units, which makes the application of these sophisticated methods possible in hospitals not having the necessary equipments and expertise.

Sperm fluorescence in situ hybridization analysis reveals normal sperm cells for 14;14 homologous male Robertsonian translocation carrier.

OBJECTIVE: To report the presence of normal sperm cells for chromosome 14 in a homologous 14;14 Robertsonian translocation carrier. DESIGN: Case report. SETTING: In vitro fertilization clinic and genetics laboratory in a private hospital. PATIENT(S): Infertile couple referred for IVF. INTERVENTION(S): Conventional cytogenetic and fluorescence in situ hybridization (FISH) techniques were used in karyotype and sperm FISH analysis. Three IVF treatments were performed, two of which included preimplantation genetic diagnosis (PGD). MAIN OUTCOME MEASURE(S): Cytogenetic analysis revealed pure 45,XY,t(14;14)(q10;q10) karyotype. Sperm FISH analysis for chromosome 14 revealed 13% normal sperm cells in the sperm sample. RESULT(S): Sperm FISH analysis revealed 13% normal sperm cells for chromosome 14 in the homologous 14;14 Robertsonian translocation carrier. The couple underwent two IVF cycles together with PGD. In the first trial there was no suitable embryo for transfer. In the second trial one normal blastocyst was transferred on day 6. However, pregnancy was not established in this second PGD cycle. CONCLUSION(S): To our knowledge, this is the first sperm FISH study revealing the presence of normal sperm in the ejaculate of a pure homologous translocation carrier. The PGD study performed for this couple is also unique in the literature.

Preimplantation genetic diagnosis (PGD) for extremes--successful birth after PGD for a consanguineous couple carrying an identical balanced reciprocal translocation.

OBJECTIVE: To report a healthy birth after preimplantation genetic diagnosis (PGD) performed for a consanguineous couple carrying an identical familial reciprocal translocation in both partners. DESIGN: Case report. SETTING: In vitro fertilization (IVF) clinic and genetic laboratory in a private hospital. PATIENT(S): Consanguineous couple carrying the same balanced reciprocal translocation: 46,XX,t(1;16)(q12;q11.2) and 46,XY,t(1;16)(q12;q11.2). INTERVENTION(S): 25 oocyte-cumulus complexes were retrieved 36 hours after human chorionic gonadotropin injection; metaphase II oocytes were fertilized by intracytoplasmic sperm injection; single blastomere biopsy was performed on 15 embryos on day 3; one embryo was found to be normal or balanced according to fluorescent in situ hybridization studies, embryo transfer was performed on day 4. MAIN OUTCOME MEASURE(S): Healthy birth of homozygous double translocation carrier twins with 46,XY,t(1;16)(q12;q11.2)mat,t(1;16)(q12;q11.2)pat karyotype. RESULT(S): Healthy monozygotic male twins were born at 36 weeks of gestation. Karyotype studies of the babies revealed that they are double translocation homozygotes: 46,XY,t(1;16)(q12;q11.2)mat,t(1;16)(q12;q11.2)pat. They are healthy and more than 4 years old later show no physical or mental abnormalities. CONCLUSION(S): To our knowledge, this is the first PGD study performed for a couple who carry the same reciprocal translocation. The twins born after this study are rare examples in the literature of healthy balanced reciprocal translocation homozygotes.