目前廣泛應用于胎兒醫學的唐氏綜合症篩查法，即結合早孕期胎兒頸項透明層的超聲檢查，及母體血清生化指標的綜合篩查法。頸項透明層是指在早孕期利用超聲檢測到的胎兒頸后的皮下積水，其作為預測胎兒異常的一項重要“軟指標，其臨床意義，尤其是與胎兒染色體異常及器官結構異常之間的關係，逐漸得到深入的認識，但其形成機制尚未明確。現在已知有一百餘種畸形及遺傳綜合征與胎兒頸項透明層增厚相關，但其染色體異常譜系，尤其是亞顯微的染色體異常仍有待明確。大部分頸項透明層增厚但核型正常的胎兒預後良好，但約3-10%的這部分胎兒會伴有畸形或出生后的神經智力發育缺陷。而傳統核型分析無法檢測到亞顯微的染色體異常，從而無法判斷這部分核型正常卻伴有缺陷的胎兒是否因為這類染色體異常而致病。 / 微陣列比較基因組雜交芯片作為檢測兒童發育遲緩者及器官結構異常原因的重要手段已廣泛應用于臨床。在染色體核型正常的胎兒中，若伴有器官結構異常的胎兒，5-12%被檢出與該畸形相關的微缺失及微重複；若僅伴有孕婦高齡或唐氏篩查高危，則微缺失及微重複檢出率約1%。 / 該課題旨在研究頸項透明層增厚但核型正常的胎兒中，染色體拷貝數變異發生的頻率及頻譜；評估微陣列比較基因組雜交芯片在協助臨床判斷胎兒預後中的作用。因此，我們開展該多中心隊列研究，通過納入449例頸項透明層厚度≧3.5 mm但正常核型胎兒的，檢測其染色體拷貝數變異，監測并記錄其圍產、產後及新生兒期情況。微陣列比較基因組雜交芯片總共檢出2.8%的異常拷貝數變異，其大小範圍為0.1 kb至18Mb。在伴有器官結構異常的胎兒組中，異常拷貝數變異檢出率達7.8%。對於頸項透明層厚度≧4.0 mm的胎兒，異常拷貝數變異檢出率可達7.3%。 / 對於頸項透明層增厚的胎兒，致病拷貝數變異暫未發現特定的頻譜。但，該研究中發現重複的致病拷貝數變異，如22號染色體長臂1區1帶的微重複或微缺失，2號染色體長臂2區2帶的微缺失。未在3號、7號、12號、13號、18號、20號、21號或Y染色體上發現與胎兒頸項透明層增厚相關的致病拷貝數變異。 / 頸項透明層增厚的胎兒79.3%預後良好；若經微陣列比較基因組雜交芯片未檢出致病拷貝數變異，則81.2%預後良好。如果僅頸項透明層增厚不伴有結構異常的胎兒，經微陣列比較基因組雜交芯片未檢出致病拷貝數變異，則93.5%預後良好。 / 綜上所述，微陣列比較基因組雜交芯片顯著提高了致病拷貝數變異的檢出率。可考慮將微陣列比較基因組雜交芯片作為頸項透明層厚度≧4.0 mm的胎兒染色體異常檢查的首要方法。對於僅頸項透明層增厚不伴有結構異常的胎兒，且經微陣列比較基因組雜交芯片未檢出致病拷貝數變異，絶大部分預後良好。對於頸項透明層增厚的胎兒，致病拷貝數變異暫未發現特定的頻譜，但發現重複出現的致病拷貝數變異。通過初步的基因本體分析及基因通路分析，神經嵴細胞的分化遷徙功能異常可作為今後研究頸項透明層增厚的病理生理機制的方向。 / Measurement of nuchal translucency (NT) has been recognized as a sensitive marker for fetal chromosomal disorders for more than a decade, and is presently used as a routine first-trimester screening test. Although over 100 abnormalities and genetic syndromes have been reported to be associated with increased NT, these associations have not been fully explored and the relevant spectrum of associated submicroscopic chromosomal abnormalities has not been sufficiently investigated. The majority of euploid fetuses with increased NT have a good outcome, but around 3-10% of fetuses present with structural or neurodevelopmental abnormalities postnatally. A range of genetic syndromes has been reported, many of which are linked to submicroscopic chromosomal abnormalities that are typically missed by conventional karyotyping. / Microarray-based comparative genomic hybridization (arrayCGH) has been applied as the first-tier diagnostic tool for the evaluation of developmental delay and structural malformations in children. In fetuses with a normal karyotype, microarray analysis revealed clinically relevant deletions or duplications in 5-12% with a structural anomaly and in about 1% of those whose indications were advanced maternal age or positive screening results. / The objectives of this study were to delineate the frequency and spectrum of pathogenic chromosome copy number variants (CNVs) among fetuses with increased NT and normal karyotype; to evaluate the role of arrayCGH to predict the prognosis of the high NT fetuses; to explore the genotype-phenotype correlations of increased NT. Therefore, a multi-centre cohort of 449 fetuses with NT ≧3.5 mm and normal karyotype were further investigated by arrayCGH. Antenatal surveillance, pregnancy outcome and paediatric follow up were documented. ArrayCGH detected abnormal CNVs in 2.8% (14 of 449) of the fetuses with high NT; the size of CNVs ranged from 0.1 kb to 18Mb. Among fetuses with major congenital abnormalities the incidence of abnormal CNV reached 7.8% (4 of 51). By adjusting the NT to ≧4.0 mm as the referral indication, 7.3% (14 of 192) of the fetuses would have abnormal arrayCGH results. The spectrum of pathogenic CNVs found associated with increased NT was diverse. However, there were recurrent ones such as the deletions or duplications at chromosomal region 22q11, and deletions in ZEB2. There was no pathogenic CNV related with increased NT found in chromosomes 3, 7, 12, 13, 18, 20, 21, or Y. The total normal outcome rate of euploid fetuses with an increased NT was 79.3%; for fetuses with normal arrayCGH results 81.2% had a normal outcome. In fetuses with isolated increased NT, normal arrayCGH results predict a favorable prognosis of 93.5%. / In conclusion, arrayCGH significantly increased the diagnostic yield of pathogenic CNVs. In clinical practice arrayCGH may be considered as the first tier investigation in fetuses with an increased NT more than 4.0 mm. In cases with an isolated increased NT with normal arrayCGH results the pregnancy outcome is likely to be favorable. The spectrum of abnormal CNVs found by arrayCGH is diverse but there are recurrent cases such as del/dup 22q11 and del ZEB2. Our preliminary gene ontology and pathway analysis showed that gene pathways related to neural crest cells may be considered as a future study for physiopathologic mechanisms of NT. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Huang, Jin. / Thesis (Ph.D.) Chinese University of Hong Kong, 2014. / Includes bibliographical references (leaves 106-120). / Abstracts also in Chinese.

La présente thèse par articles aborde différentes facettes du dépistage prénatal de certaines aneuploïdies au premier trimestre de la grossesse. L’introduction retrace l’historique du dépistage prénatal et énonce les différents marqueurs biochimiques et échographiques associés aux aneuploïdies. La première publication démontre que le tabagisme maternel abaisse significativement les niveaux sanguins maternels de PAPP-A et de la fraction libre de la β-hCG et augmente significativement la clarté nucale, confirmant la nécessité de contrôler cette co-variable dans le calcul de risque final, du moins pour la trisomie 18. Le deuxième article identifie des seuils de clarté nucale au-delà desquels la biochimie génétique n’apporte aucune valeur additionnelle au dépistage prénatal de la trisomie 21 et de la trisomie 18. Pour les fœtus avec clarté nucale supérieure aux seuils établis, un diagnostic prénatal intrusif devrait être offert sans délai. Le troisième et dernier article porte sur la première détermination des niveaux plasmatiques maternels de la protéine FLRG (follistatin-related gene) au premier trimestre de grossesse et sur son rôle potentiel à titre de marqueur biochimique dans le dépistage prénatal de la trisomie 21. Bien que détectables, les niveaux plasmatiques maternels de FLRG ne sont pas significativement altérés en présence de fœtus avec syndrome de Down. Dans la discussion générale, les trois articles sont abordés sous un angle plus spécifique au Québec. Des données complémentaires et originales y sont présentées. Une discussion sur l’évolution future du dépistage prénatal est entamée et des axes de recherche sont proposés. / In this thesis by articles, we explore different facets of first trimester prenatal screening of aneuploidy. Introduction retraces the origin of prenatal screening and enunciates current biochemical and ultrasound markers associated with aneuploidy. In the first article, impact of maternal smoking on first-trimester prenatal screening results is assessed for Down syndrome and trisomy 18. Both maternal blood levels of PAPP-A and free β-hCG are significantly decreased by maternal smoking while fetal nuchal translucency (NT) thickness is significantly increased. Without adjustment, this results in an increase of false positives, at least for trisomy 18. Based on these results, adjustment for smoking should be mandatory in first-trimester prenatal screening. In the second article, we identify NT threshold values above which biochemical screening provides no additional benefit. In pregnancies in which NT is above the proposed upper cut-offs, invasive prenatal screening should be offered without undue delay. In the third and last article, maternal plasma levels of follistatin- related gene protein (FLRG) are determined for the first time in first trimester of pregnancy. Its potential role as a new marker for Down syndrome is assessed. Although FLRG can be successfully detected in maternal plasma, its levels are not significantly altered by the presence of Down syndrome fetuses. In the general discussion, articles are mainly addressed under a Quebec standpoint. Additional and complementary original data are presented and different clinical research avenues are proposed.

La présente thèse par articles aborde différentes facettes du dépistage prénatal de certaines aneuploïdies au premier trimestre de la grossesse. L’introduction retrace l’historique du dépistage prénatal et énonce les différents marqueurs biochimiques et échographiques associés aux aneuploïdies. La première publication démontre que le tabagisme maternel abaisse significativement les niveaux sanguins maternels de PAPP-A et de la fraction libre de la β-hCG et augmente significativement la clarté nucale, confirmant la nécessité de contrôler cette co-variable dans le calcul de risque final, du moins pour la trisomie 18. Le deuxième article identifie des seuils de clarté nucale au-delà desquels la biochimie génétique n’apporte aucune valeur additionnelle au dépistage prénatal de la trisomie 21 et de la trisomie 18. Pour les fœtus avec clarté nucale supérieure aux seuils établis, un diagnostic prénatal intrusif devrait être offert sans délai. Le troisième et dernier article porte sur la première détermination des niveaux plasmatiques maternels de la protéine FLRG (follistatin-related gene) au premier trimestre de grossesse et sur son rôle potentiel à titre de marqueur biochimique dans le dépistage prénatal de la trisomie 21. Bien que détectables, les niveaux plasmatiques maternels de FLRG ne sont pas significativement altérés en présence de fœtus avec syndrome de Down. Dans la discussion générale, les trois articles sont abordés sous un angle plus spécifique au Québec. Des données complémentaires et originales y sont présentées. Une discussion sur l’évolution future du dépistage prénatal est entamée et des axes de recherche sont proposés. / In this thesis by articles, we explore different facets of first trimester prenatal screening of aneuploidy. Introduction retraces the origin of prenatal screening and enunciates current biochemical and ultrasound markers associated with aneuploidy. In the first article, impact of maternal smoking on first-trimester prenatal screening results is assessed for Down syndrome and trisomy 18. Both maternal blood levels of PAPP-A and free β-hCG are significantly decreased by maternal smoking while fetal nuchal translucency (NT) thickness is significantly increased. Without adjustment, this results in an increase of false positives, at least for trisomy 18. Based on these results, adjustment for smoking should be mandatory in first-trimester prenatal screening. In the second article, we identify NT threshold values above which biochemical screening provides no additional benefit. In pregnancies in which NT is above the proposed upper cut-offs, invasive prenatal screening should be offered without undue delay. In the third and last article, maternal plasma levels of follistatin- related gene protein (FLRG) are determined for the first time in first trimester of pregnancy. Its potential role as a new marker for Down syndrome is assessed. Although FLRG can be successfully detected in maternal plasma, its levels are not significantly altered by the presence of Down syndrome fetuses. In the general discussion, articles are mainly addressed under a Quebec standpoint. Additional and complementary original data are presented and different clinical research avenues are proposed.

Abstract The purpose of this study was to evaluate extended first trimester screening for severe chromosomal disorders and adverse pregnancy outcomes in singleton pregnancies among the general population in Finland. Maternal serum biochemical markers, pregnancy associated plasma protein-A (PAPP-A) and free beta human chorionic gonadotropin (fβ-hCG), and fetal nuchal translucency (NT) thickness were measured during the gestational weeks 8+0–13+6. A computerized risk figure program was used to calculate an individual risk figure for chromosomal disorders. It was investigated whether the screening parameter, PAPP-A, is associated with adverse pregnancy outcomes. The prevalence of Down syndrome (DS) cases in Finland during the years 2002–2006 was 1:364 (N=795). The proportion of women aged 35 years or older increased from 5–10% in the years 1980–1990 to 19.1% during the study period. Most DS cases (61.1%) presented in that age group. The first trimester combined screening for Down syndrome yielded a detection rate (DR) of 81.9% for a 4.3% false positive rate (FPR). The performance was evaluated among 76949 voluntary women during the study period of 01.05.2002–31.12.2008. There were 188 cases of DS. The screening worked better among the older women. The number of invasive procedures needed to detect one case of DS was higher among the younger women. Adding specific algorithms for screening of other chromosomal abnormalities yielded DR of 74.0% for trisomy 18 (T18) and 54.5% for trisomy 13 (T13) with an additional increase of 0.3% FPR. For chromosomal abnormalities other than T18 and T13, the specific algorithms did not improve the screening performance. Low first trimester maternal serum levels of PAPP-A (≤0.30 MoM) were significantly associated with small for gestational age (SGA) newborns and stillbirths (SBs). The combined screening method for DS works well in practice and has been standardized in Finland. In screening for trisomies 18 and 13 a specific algorithm is reasonable. Low first trimester levels of PAPP-A could be used as an independent marker for pregnancies at high risk for SGA babies and SBs. / Tiivistelmä Tutkimuksen tarkoituksena oli arvioida laajennetun ensimmäisen raskauskolmanneksen kromosomipoikkeavuuksien seulonnan toimivuutta yksisikiöisissä raskauksissa suomalaisessa normaaliväestössä. Äidin seerumin biokemialliset merkkiaineet, raskauteen liittyvä valkuaisaine A (PAPP-A) ja raskaushormoni (fβ-hCG) sekä sikiön niskaturvotus mitattiin raskausviikoilla 8+0–13+6. Yksilöllinen riskiluku kromosomipoikkeavuuksille laskettiin käyttäen tietokoneen riskinlaskentaohjelmaa. Seulonnan merkkiaineen, PAPP-A:n, matalien pitoisuuksien yhteyttä epäsuotuisiin raskauden lopputuloksiin tutkittiin. Downin oireyhtymän esiintyvyys Suomessa oli 1:364 (N=795) vuosina 2002–2006. 35-vuotiaiden tai sitä vanhempien naisten osuus oli tutkimusaikana 19.1 %, mikä on huomattavasti suurempi kuin vuosien 1980–1990: 5–10 %. Näiden naisten sikiöiden joukosta löytyi suurin osa Down oireyhtymistä (61.1 %). Ensimmäisen raskauskolmanneksen yhdistelmäseulonnan toimivuutta tutkittiin aikana 01.05.2002–31.12.2008. Tutkimukseen osallistui 76 949 vapaaehtoista naista. Joukossa oli 188 Downin oireyhtymätapausta. Seulonnan herkkyys Downin oireyhtymälle oli 81.9 % ja tarkkuus 4.3 %. Seulonta toimi parhaiten vanhempien naisten joukossa. Niiden kajoavien toimenpiteiden määrä, jotka tarvittiin yhden Down-sikiön löytämiseksi, oli suurempi nuorten naisten joukossa. Tutkimuksessa Downin oireyhtymän algoritmiin lisättiin spesifiset algoritmit trisomioille 18 ja 13, jolloin saavutettiin 74.0 %:n herkkyys trisomialle 18 ja 54.5 %:n herkkyys trisomialle 13. Väärien positiivisten seulontatulosten määrä kasvoi 0.3  %:n verran. Seulonnan toimivuus muiden kromosomipoikkeavuuksien joukossa ei parantunut spesifisten algoritmien avulla. Lisäksi matalan PAPP-A-pitoisuuden yhteys pienipainoisuuten ja kuolleena syntyneisyyteen oli tilastollisesti merkittävä. Tutkimus osoitti, että esimmäisen raskauskolmanneksen yhdistelmäseulonta toimii hyvin käytännössä. Trisomioiden 18 ja 13 seulonnassa spesifisten algoritmien käyttö on järkevää. Matalaa ensimmäisen raskauskolmanneksen PAPP-A-arvoa voitaisiin käyttää itsenäisenä riskimerkkiaineena raskauksille, joissa pienipainoisuuden ja kuolleena syntymisen riski on kohonnut.