Genetically isolated populations
Implications for genetic care
Mathijssen, I.B.
Publication date
2018
Document Version
Other version
License
Other
Link to publication
Citation for published version (APA):
Mathijssen, I. B. (2018). Genetically isolated populations: Implications for genetic care.
General rights
It is not permitted to download or to forward/distribute the text or part of it without the consent of the author(s) and/or copyright holder(s), other than for strictly personal, individual use, unless the work is under an open content license (like Creative Commons).
Disclaimer/Complaints regulations
If you believe that digital publication of certain material infringes any of your rights or (privacy) interests, please let the Library know, stating your reasons. In case of a legitimate complaint, the Library will make the material inaccessible and/or remove it from the website. Please Ask the Library: https://uba.uva.nl/en/contact, or a letter to: Library of the University of Amsterdam, Secretariat, Singel 425, 1012 WP Amsterdam, The Netherlands. You will be contacted as soon as possible.
DISCUSSION
Genetically isolated populations provide unique opportunities for many different genetic investigations. Examples are: identification of genes that underlie Mendelian (autosomal recessive) disorders; identification of susceptibility genes predisposing to complex disorders; studying penetrance, phenotypic variation, and modifying factors underlying the clinical variability of Mendelian disorders; and implementation and evaluation of genetic carrier screening programs. The main objective of this thesis is to demonstrate these various opportunities in a young genetically isolated population in the Netherlands.
In this chapter, the main findings of this thesis will be presented and discussed in the context of the scientific literature. The chapter concludes with future perspectives for research and clinical care in genetically isolated populations.
Identification of genes that underlie autosomal recessive disorders
Genetically isolated populations are very suitable for identifying new genes because of the reduced genetic heterogeneity. A variety of strategies can be used to identify these genes. We used an approach of homozygosity mapping1 followed by Sanger sequencing of candidate genes to identify new founder mutations of two different disorders in the genetically isolated population under study (chapter 2 and 3).
Identification of new founder mutation (FADS) and reproducibility
The first disorder is fetal akinesia deformation sequence (FADS) (MIM 208150), a lethal disorder characterized by progressively reduced fetal movements, joint contractures, and severe pulmonary hypoplasia (chapter 2). We identified a homozygous missense muta-tion in the MUSK gene as the cause of FADS in this genetically isolated populamuta-tion.2 The MUSK gene is essential for the development of the neuromuscular junction. Muta-tions in this gene had been described before in patients with congenital myasthenic syndrome (CMS), but not in lethal FADS. However it was already shown that muta-tions in other genes encoding neuromuscular synaptogenesis can cause phenotypes with variable severity, ranging from CMS to FADS. With the identification of MUSK as a novel cause of lethal FADS, this gene can be included in this row of genes. To confirm our finding, we tested a cohort of 26 unsolved cases with FADS who did not come from the genetically isolated population. However, no likely pathogenic mutations in MUSK were identified.
Findings in genetically isolated populations may not be reproducible in the general population. This may especially be a problem in association studies for complex trait
9
gene identification,3,4 but also for identification of genes that underlie Mendelian disor-ders. Even if found in the general population, these mutations may only explain a small subset of the genetic causes of the disorder. This is for example the case in primary ciliary dyskinesia (PCD) (MIM 615067)5 and osteogenesis imperfecta type IIB/III (OI) (MIM 610682)6 in the genetically isolated population under study.
Shortly after the publication of our article,2 another article was published about a family with recurrent pregnancies with lethal FADS which was caused by a homozygous frameshift mutation in MUSK.7 This confirms that also outside the genetically isolated population under study mutations in MUSK are a (probably) rare cause of FADS.
Identification of new founder mutation (XX-GD) and genetic heterogeneity
The second disorder is XX female gonadal dysgenesis (XX-GD), a genetically heteroge-neous disorder characterized by primary amenorrhea, lack of secondary sexual character-istics, streak gonads, hypoplastic uterus, and hypergonadotropic hypogonadism (chapter
3). More than 10 patients with XX-GD are known in the genetically isolated population. About half of these patients also suffer from sensorineural deafness. The combination of XX-GD and sensorineural deafness is known as Perrault syndrome (PS) (MIM 233400). Whole exome sequencing (WES) in three patients with symptoms of PS initially showed no variants present in all three patients (personal communication), however two patients were homozygous for the c.707A>T, p.(Asn236Ile) mutation in the ERAL1 gene.8 Also a third female PS-patient and a male patient with sensorineural deafness turned out to be homozygous for this missense mutation. Sanger sequencing of five patients with XX-GD without sensorineural deafness in this population did not reveal the founder mutation in ERAL1, suggesting genetic heterogeneity for XX-GD in this population. We decided to perform homozygosity mapping of four of the patients with XX-GD without deaf-ness, followed by Sanger sequencing of the candidate gene PSMC3IP. All five cases with XX-GD without deafness were homozygous for the c.74G>C, p.(Arg25Pro) missense mutation in PSMC3IP. Interestingly, also the case with presumed PS (with XX-GD and sensorineural deafness) turned out to be homozygous for the PSMC3IP founder muta-tion, suggesting the hearing loss in this patient has a separate cause. This case shows that also in genetically isolated populations genetic heterogeneity exists.
Genetic heterogeneity was experienced before in this genetically isolated population. Several patients with autosomal recessive retinitis pigmentosa (RP) are known in this population, and initially it was assumed that it was caused by a single type of RP.9,10 However, clinical heterogeneity was observed, with the majority of the patients having RP with preserved para-arteriolar retinal pigment epithelium (PPRPE), while this
phenotype was not found in the remaining RP-patients.11,12 In the patients with PPRPE, a homozygous mutation in CRB1 was identified, causing RP12 (OMIM 600105).13 In the remaining patients three other causes of RP were identified (Table 1).14
Also in other genetically isolated populations both non-allelic15 as well as allelic hetero-geneity16 have been demonstrated, which can complicate the genetic mapping of some disorders. However, despite this caution, the chance of experiencing allelic and non-allelic heterogeneity in genetically isolated populations is reduced and these populations are very useful for identifying new genes.
Phenotypic variation
In outbred populations it may be difficult to study phenotypic variation in patients with the same homozygous mutation because extended pedigrees and larger series of patients with the same mutation are rare and non-genetic variance (environmental and cultural factors) influencing the phenotype may be more evident.
Genetically isolated populations, however, provide a unique opportunity for studying phenotypic variation, penetrance, and modifying factors underlying the clinical vari-ability of autosomal recessive disorders. The majority of the patients are homozygous for the same founder mutation, tend to show reduced non-genetic variance, and frequently it is easy to re-contact patients for follow-up studies as many of the inhabitants stay within the community.
In the genetically isolated population under study we studied the long-term clinical course and variability of 30 patients with retinitis pigmentosa type 12 (RP12) due to a homozygous founder mutation in the CRB1 gene (chapter 4). The age of diagnosis and the clinical course (in terms of visual acuity, visual fields, and ophthalmic findings) showed marked interindividual variability. This supports the hypothesis that the pheno-type of patients with CRB1 mutations is modulated by other factors.14
In this genetically isolated population, phenotypic variation was also studied in several patients with pseudoxanthoma elasticum (PXE) caused by a homozygous founder muta-tion in ABCC6.17 Also, considerable interindividual phenotypic variability was demon-strated for this disorder.
As in both genetic disorders, RP12 and PXE, the interindividual difference between the individuals with the same homozygous mutation is very high, studying geno-type-phenotype correlations for these disorders are expected to be fruitless. Both genetic disorders can be used for unraveling genetic and environmental factors modifying the phenotypes.
9
Knowledge of founder mutation enhances better care
Each genetically isolated population shows a unique profile of rare disease alleles, with some genetic disorders (much) more prevalent compared to the general population, while others are less prevalent.
In the genetically isolated population under study, founder mutations for 17 Mendelian disorders are currently known (Table 1). The carrier frequencies of the autosomal reces-sive disorders are generally (very) high, ranging from about 2 to 14%.
In most of the genetically isolated populations in the Netherlands just zero to a handful of Mendelian disorders have been identified,27,28 being much less than in the genetically isolated population under study. There are several possible reasons for this difference; the founder individuals in the other populations were carrier of none or only a few genetic disorders, the founder population is less isolated and/or younger and/or larger, insufficient knowledge about the disorders is present in the population, the founder population is assumedly not as cooperative in genetic research (e.g. because of religious barriers), and insufficient recognition of patterns of disorders in the population. Recog-nition of these (patterns of) disorders is greatly enhanced by drawing pedigrees during genetic counseling in members of the genetically isolated population. The genetically isolated population under study is very cooperative in genetic research.
Knowing which disorders are more prevalent in a genetically isolated population, including their specific genes and mutations, is important for making a rapid and correct (differential) diagnosis by clinicians, for genetic counseling to help individuals, couples and families in the population to understand and adapt to the medical, psychological, familial and reproductive implications of the genetic disorder(s),29 and for research in these populations. For some genetically isolated populations (online) databases with an overview of genetic disorders (much) more prevalent in these populations are available. Examples are the Amish, Mennonite, and Hutterite Genetic Disorder Database (www. biochemgenetics.ca/plainpeople),30 the Finnish Disease Database (www.findis.org),31 and the Israeli National Genetic Database (www.goldenhelix.org/israeli).32 Currently, such a database is not available in the Netherlands and it is debated whether we should develop it. Besides the above mentioned advantages, there may be disadvantages, such as stigmatization. This may be a reason not to develop an open access database but instead a database only accessible to health care providers.
Carrier screening in genetically isolated populations
In the genetically isolated population under study four severe recessive genetic disorders occur at relatively high frequency; pontocerebellar hypoplasia type 2 (PCH2) (MIM 277470), fetal akinesia deformation sequence (FADS), rhizomelic chondrodysplasia punctata type 1 (RCDP1) (MIM 215100), and osteogenesis imperfecta (OI) type IIB/III. Children with these disorders suffer significant morbidity and have a severely reduced lifespan. An estimated 2 to 4 of the 250 children born annually are affected with one of these disorders. In September 2012 we started an outpatient clinic in the genetically isolated population offering carrier screening for these four disorders, aimed
9
at couples planning a pregnancy (chapter 5).
Genetic carrier screening showed a very high carrier frequency of 33% of the four tested disorders. Not only in individuals with a positive family history for one of the disorders, but also in individuals with a negative family history, the carrier frequency was high (57.8% versus 25.8%).18 In the first year, four PCH2 carrier couples were identified. Using cascade genetic carrier testing, in which carrier testing is targeted to close rela-tives and partners of previously identified patients and carriers, the carriers and carrier couples without a positive family history would not have been identified.
Targeted carrier screening programs for multiple disorders have been introduced in a few other genetically isolated populations worldwide, for example in people of Eastern European Jewish (Ashkenazi) (AJ) descent,33 in different isolated (mainly Arab and Druze) communities in Israel34-36 and in the Saguenay-Lac-Saint-Jean region of Quebec, Canada.37 Also in these populations, carrier frequencies are high and these programs are quite successful in terms of providing couples with meaningful options for autonomous reproductive choice (the primary aim of carrier screening).38 Targeted carrier screening in the genetically isolated population under study, but also in other genetically isolated populations with a high prevalence of specific disorders, is thus an effective method to identify carriers and is likely to be more effective compared to cascade genetic testing. In the genetically isolated population under study, also other (non-lethal) disorders are highly frequent, for example phenylketonuria (PKU) (MIM 261600), primary ciliary dyskinesia (PCD) (MIM 615067), and retinitis pigmentosa (RP) (see Table 1). If there is a positive family history for (one of) these other disorders, we offer further genetic counseling for the specific disorder(s). Because of the less severe nature of these disor-ders, therapeutic options in some of them, and the (presumably) different reproductive consequences, it is preferred not to include these less severe disorders in the test panel with the severe disorders, but instead to offer one or more separate test panels for which the couples can give separate informed consent.39,40
It is important to carefully consider if and how to offer carrier screening for milder disor-ders in genetically isolated populations and in the general population. In the AJ popula-tion carrier screening for Gaucher disease (GD) was included in the test panel because it is one of the most prevalent recessive disorders in this population, for which testing is simple and the test sensitivity is high.41 However, the disorder is frequently mild or asymptomatic and therapeutic options (enzyme replacement therapy) exists. It is therefore ethically ques-tionable to systematically carry out carrier screening for this disorder.41,42
We recently identified a relative frequent founder mutation for MUTYH-associated polyposis coli (MAP) (OMIM 608456) in the genetically isolated population under study. Individuals who inherit MUTYH mutations from both parents (biallelic) have a colorectal cancer (CRC) risk of about 43% at age 60 and the life-time risk is assumed to be close to 100% in the absence of timely surveillance.43 The lifetime risk for duodenal cancer is about 4%.43 Surveillance by colonoscopy and gastro-duodenoscopy is advised in MAP patients. In the genetically isolated population under study, the carrier frequency of MAP is 9.8% (compared to 1-2% in the general population44,45). The chance of being homozygous for this mutation in this population is about 1 in 400, being much more frequent than in the general population. The first steps have been taken in informing at-risk families and offering genetic testing for MAP in the genetically isolated popula-tion under study (see Future perspectives).
In the AJ population there is already experience with carrier screening for late-onset onco-genetic disorders. In this population, three breast-cancer founder mutations (two BRCA1-mutations and one BRCA2-mutation) are more prevalent than in the general population. It is estimated that over two percent of the AJ population is a carrier of one of these mutations.46 Genetic testing of these three founder mutations is offered on a research basis, in for example Israel, the UK, and Canada, to individuals of AJ descent, regardless of personal or family history of cancer.47-49 Most women with mutations would not have been qualified for genetic testing based on personal and/or family history of cancer.50 It has been argued that genetic testing of the general population of Jewish women is justified.50 In these studies, carrier screening for founder mutations in BRCA1 and BRCA2 is offered separately from the carrier screening for the recessive founder mutations.
Expanded carrier screening and genetically isolated populations
Meanwhile, technological advances have enabled the development and offer of precon-ception expanded carrier screening (ECS) in which couples without an a priori increased risk of having a child with a genetic disorder can be screened for several (hundreds of) disorders simultaneously.51,52 An increasing number of mainly commercial laborato-ries offer these screening panels.39,53 The existing (commercial) ECS panels may be less suitable for inhabitants of genetically isolated populations, as the carrier frequencies of several autosomal recessive disorders in these populations can be very skewed from nationwide or worldwide carrier frequencies.
We made an inventory of six Dutch genetically isolated populations and their auto-somal recessive founder mutations, and investigated whether the founder mutations
9
were covered in the (preconception) expanded carrier screening tests of carrier screening providers (chapter 8). The great majority of these founder mutations are not covered in the ECS panels of the five selected providers.27 This also applies to most of the founder mutations present in genetically isolated populations in other countries. Offering a (commercial) routine ECS panel to inhabitants (of the majority) of the genetically isolated populations is not appropriate because it may give false reassurance to couples with an increased risk of having a child with a founder mutation related disorder they are not being tested for. It is important that descendants from genetically isolated popu-lations are aware that a (commercial) routine ECS panel may not be appropriate and may give false reassurance. Currently, this information is not mentioned by expanded carrier test providers.
Ideally, a customized ECS test will be developed for each country/region in which both country/region-specific mutations as well as genetic isolate-specific founder mutations are present. This approach may also reduce potential stigmatization of genetically isolated populations, while specific mutations still are included. However, preventing high healthcare costs may be an important reason against a country/region wide screening test instead of a genetic-isolate wide screening test.54 Also, the preferences of the target population are important in this context.54
In May 2016 the Academic Medical Center (AMC) Amsterdam started a non-profit offer of expanded carrier screening for 50 severe recessive disorders (www.dragerschaps-test.nl). Most of the severe disorders currently known in Dutch genetically isolated populations are included.
It is expected that in the near future WES or even whole genome sequencing (WGS) will be used for carrier screening. The first steps have already been taken.55 A great advantage is that all known disease genes can be screened, including very rare disease genes prevalent in genetically isolated populations and in consanguineous couples.56 However, correct interpretation of test-results when using WES or WGS is complex and it is very important that only variants known to affect function (and not variants of unknown clinical significance) are reported.52 Also, the identification of carrier couples for mild disorders which are unlikely to alter reproductive plans is an important point to take into consideration.55,57
Recent developments and results of carrier screening in the
genetically isolated population under study:
• In June 2014 the carrier testing panel offered in the outpatient clinic was expanded from four to five recessive disorders by adding a founder mutation in the POLG
gene to the panel after the discovery of a couple of patients with this disorder in the population. Mutations in the POLG gene cause a continuum of phenotypes with variable severity and age of onset. We expect that homozygosity for the founder mutation c.2243G>A, p.(Trp748Ser) is lethal in the early embryonic period, based on the expected deleterious effect of the mutation and the absence of individuals with homozygosity for this mutation. However, compound homozygosity of the above mentioned founder mutation combined with another mutation may cause a phenotype with intellectual disability, seizures, ataxia, liver dysfunction, neuropathy and/or progressive external ophthalmoplegia.
• After five years of carrier screening, in total 12 carrier couples for one of the four diseases have been identified; five for PCH2, three for FADS, two for OI, and two for RCDP1. The chance of being a carrier couple for one of the four disorders is about 4%. Beside this, eight carrier couples for POLG have been identified (in five of them both partners are carrier of the above mentioned POLG founder mutation, in three of them one of the partners is carrier of the founder mutation and the other of another mutation).
• In the first year of the outpatient clinic the majority (89%) of the couples decided to test simultaneously. However, five years later the opposite is the case as most couples prefer sequential testing, in which one of the partners is tested first and DNA testing is only performed on the other partner if the first one turns out to be a carrier. The main reason for this is not to pay the deductible excess (obligatory deductible excess was 385 euros in 2017) of both partners.
• Since May 2016, some couples (n=6 until November 2016) from the genetically isolated population under study asked for the AMC ECS test instead of the targeted screening test. Only carriers for one of the five disorders also present in the targeted screening test were identified. This is not surprising, as in the genetically isolated popu-lation no patients with the other disorders included in the screening panel are known. • As mentioned above, the first steps have been taken in implementing genetic testing
for MAP in the genetically isolated population under study.
Evaluation of the preconception carrier screening offer
When introducing carrier screening, it is important to investigate the (potential) beneficial and harmful implications of the screening as well as preferences regarding genetic coun-seling. Many studies have been published about the implementation of carrier screening for a single disorder. In general, these studies demonstrated that screening is well received by the participants without major adverse psychological effects, and showed that the
9
participants intended to base their reproductive decisions on the test results.52 However, studies about carrier screening for multiple disorders simultaneously are scarce.37,58-60 Evaluating the screening offer in the genetically isolated population under study provides lessons for optimizing the screening in this population as well as for ECS in the general population, although not all results can be generalized because of social and cultural differences between the genetically isolated population and the general population. The carrier screening for four disorders offered in the outpatient clinic was evaluated by administering questionnaires (both attendees and non-attendees) and conducting semi-structured interviews with carrier couples (chapter 6).
The attendees were highly satisfied and would recommend screening to others. All identified carrier couples made reproductive decisions based on their results, such as refraining from having children, prenatal diagnosis (PND), and preimplantation genetic diagnosis (PGD). No major adverse psychological effects and no feelings of stigmatiza-tion were reported (Mathijssen et al, in press).
We have also identified challenges when offering carrier screening. Although the majority of the people were highly familiar with the genetic disorders and carrier testing and knowledge was high, some people wrongly mentioned an increased risk of having an affected child if both partners are carriers of different recessive disorders. It is impor-tant to emphasize this topic in the counseling of carrier screening for multiple disorders. Most participants recalled their test results correctly, but two couples reported being carrier of another disorder than reported to them. It can be expected that, when expanding the number of disorders screened for and more carriers being identified, recall of test results will become more difficult, and thus requires explicit attention in information provision and counseling.
The majority of the participants stated that couples should always have pre-test consul-tation, and did not prefer a GP over a genetic counselor. In a focus group study with forty US genetics professionals, there was consensus that ECS should be accompanied by pre- and post-test genetic counseling, preferably by a clinician with expertise in communicating genetic information.61 In a Dutch online survey, most potential users of ECS also preferred face-to-face consultation but the majority preferred the test to be offered via their GP, probably due to the strong primary care structure in the Neth-erlands.62 In the genetically isolated population under study, counseling of about 200 individuals a year is achievable, but when considering a nation-wide ECS, face-to-face counseling with a genetic counselor is impossible because there are not enough genetic
professionals.51 Counseling by non-genetic professionals (e.g. GPs), more personalized information, using for instance interactive computerized information/apps, and easily accessible telephone contact with a genetic professional, could be a solution.52,63
Most individuals in the genetically isolated population were informed about the existence of the carrier screening and the outpatient clinic by close influencers (family/ friends). When introducing ECS in other populations, it might be worth-while to use these influencers to raise awareness about the screening-test offer. In the general population, however, community support is less evident as there is no specific community with which people can identify themselves.38
Although not all results of the evaluation of carrier screening in this genetically isolated population may be generalizable to the ECS in the general population, some of the recommendations may be useful. Important in this context is that we have found no evidence that screening for multiple disorders will cause major adverse psychological effects. Moreover, the recall and interpretation of test results may become more chal-lenging for participants in multi-disorder carrier screening, which requires explicit attention in information provision and counseling.
Factors contributing to successful implementation of carrier screening
The implementation of the carrier screening in the genetically isolated population under study was proved to be very successful. We can learn from this and other implemented initiatives by identifying critical factors involved in this successes (chapter 7).
Our study identified several critical factors, from a user perspective, in the genetically isolated population under study that contribute to successful implementation:38
• Familiarity. The familiarity with the genetic disorders and carrier screening was high compared to the general population. Of the respondents, 62% knew someone with a severe genetic disorder and 82% had heard about carrier screening.
• Perceived benefits & perceived social barriers. The respondents were highly positive
about carrier screening in their community and perceive high personal benefits and low social barriers (e.g. stigmatization) to carrier screening.
• Acceptance of reproductive options. The respondents showed a relative high acceptance of reproductive options, such as PND and PGD, and also termination of pregnancy in case of an affected fetus was considered acceptable.
• Community support. Although the initial demand for carrier screening did not entirely come from the community itself, the high social adhesion of the community facili-tated the implementation process after its introduction by health care professionals.
9
These above mentioned important factors were also present in the AJ population being another high-risk population.38
For population-based ECS, the above mentioned factors for successful implementation may be less evident as the general population is expected to be less familiar with genetic disorders and carrier screening, the perceived risk of being a carrier is low, and people are expected to be unaware of the possible benefits of the carrier screening. To ensure successful implementation of population-based ECS, efforts should be made to increase knowledge about genetic disorders, create awareness, and address personal benefits of carrier screening in a non-directive way.
(Ethical) challenges
Although genetically isolated populations provide unique opportunities for many different genetic investigations, several potential (ethical) challenges exist. Some of these issues may especially be important in genetically isolated populations compared to the general population.
•
Unequal access to genetic care
The unique opportunities for many different genetic investigations in genetically isolated populations may increase genetic research and the development of carrier screening programs in these populations. However, this may limit attention to other populations or the general population. This is for example the case in Tay-Sachs disease, in which research is almost exclusively focused on the AJ population, leaving other groups including French Canadians who have a high prevalence of the disease, less well served.64 Research attention to BRCA1/2 in the AJ population may well generate similar disparities.64
Furthermore, in many countries, including the Netherlands, carrier screening is only offered to subpopulations with a known increased risk of being a carrier of a selected group of disorders, causing unequal access to these services. Offering ECS to all individuals may result in equitable application of genomic technology.65 However, several interviewed stake-holders in a Dutch study by Van der Hout et al.66 mention that ECS might hinder rather than promote equity of access to carrier screening as the specific needs of people belonging to a specific risk group might be lost from sight when offering ECS.
•
Discrimination and stigmatization
Most studies have revealed no predominant feelings of stigmatization among carriers,52 although in some studies concerns exist about social stigmatization of
carriers (e.g. in the AJ population)67 and self-stigmatization of carriers expressed by poorer health perception (e.g. in CF carrier screening).68
The genetic cause of PCH2 has been discovered in the genetically isolated population under study. Several individuals, including inhabitants of the genetically isolated popula-tion and health care providers, call the disorder after the name of the village. This should be avoided as this suggests that the disorder is exclusive to the genetically isolated popula-tion under study, which may increase stigmatizapopula-tion. The choice of words laboratories and clinicians use to describe carrier screening test panels in de AJ population (such as “Ashkenazi Jewish Genetic Disease Panel”) raises similar concerns.69 However, it is diffi-cult to think of another name that covers the content of the test.
Beside this, media attention about the carrier screening offer(s) in genetically isolated population(s) may trigger stigmatization. The reasons for health care providers to participate in media interviews may be to inform the genetically isolated population as well as the general population about the chance of having a child with an auto-somal recessive disorder including the opportunity of carrier screening. However, journalists often filter out the comments about the general population and focus on the more intriguing genetically isolated population itself (see Appendix).
Due to carrier screening, the number of children born with the screened auto-somal recessive disorders may reduce and these disorders may be increasingly seen as preventable disorders.66 This may cause disability-based stigmatization by making the society less tolerant of affected individuals and their parents.70
A test targeted to a particular subpopulation may cause population/ethnicity-based stig-matization. Offering population-based ECS might decrease this possible stigmatization by offering a ‘universal’ test rather than targeted to a particular subpopulation.52,66 •
Undue pressure on individual choice
Individuals may feel an obligation to participate in genetic research and/or may feel a pressure to participate in carrier screening.71,72 This may especially be a concern in socially tight communities.71
FUTURE PERSPECTIVES
Research perspectives
Research in the genetically isolated population under study has proved to be successful and this population offers several opportunities for further research. For each research
9
opportunity it is however very important to carefully consider the potential beneficial and harmful aspects for the population. Some of these potential harmful aspects are discussed in the Ethical considerations-section. Other aspects are incidental findings73 and concerns in the participants about sharing genetic research data.74
Suggestions for further research are:
• Although founder mutations for 17 Mendelian disorders are currently known in the genetically isolated population under study, founder mutations for several other disorders still have to be identified. We are currently searching for Mendelian causes of intellectual disability and hereditary cardiac disorders, among others.
• For several disorders in the genetically isolated population, considerable phenotypic variation in individuals with the same founder mutation was shown. This popula-tion offers a unique opportunity for unraveling genetic and environmental factors modifying the phenotypes.
• As genetically isolated populations are also very valuable for identification of suscep-tibility genes predisposing to complex disorders (see introduction of this thesis),75,76 also the genetically isolated population under study gives an opportunity for complex trait gene identification.
• Genetically isolated populations may be more suitable for designing induced pluripo-tent stem cell (iPS) models compared to the general population because of the higher chance of finding multiple individuals with (the same) long ROHs that include disease risk loci. After reprogramming adult cells into pluripotent stem cells, these cells can be converted to differentiated cells (such as neurons), giving the oppor-tunity to study the underlying functional mechanisms contributing to disease risk and for drug development.77 The first steps for designing iPS cell models for brain related-disorders (e.g. autism) in the genetically isolated population under study have already been taken.
• With the identification of a relative frequent founder mutation for MUTYH-associ-ated polyposis coli (MAP) in the genetically isolMUTYH-associ-ated population under study, questions arise about carrier testing for this disorder in the population. (How) should we imple-ment carrier screening for this late-onset oncogenetic disorder? What are the beneficial and harmful aspects of carrier testing for late-onset disorders from a user perspective? Should we combine this testing with the offer of preconception carrier screening for severe recessive disorders? These questions are not only important for the genetically isolated population itself but will also provide lessons for the implementation of carrier screening for late-onset and (possibly) treatable disorders in the general population.
With the increased mobility of humans, genetically isolated populations become less isolated and eventually will disappear. Performing genetic research in these populations should therefore not be postponed.
• New genetic testing technologies and decreasing costs will inevitably cause expan-sion of the use of ECS from selected high-risk populations to several other patient groups or even the general population. The question is whether we should offer ECS in high-value pregnancies, for example in couples undergoing IVF/ICSI/PGD, and to gamete donors or to the general population. Also, the question is which disorders are appropriate to be included in the screening panel based on clear and transparent criteria (e.g. severity of the disorder, age of onset, availability of treatment, carrier frequency). Further studies are needed to investigate the preferences of the couples/ donors/recipients and the implications for the health care system, including the availability of trained (genetic) counselors, and (down-stream) costs.
Future clinical perspectives
It is important to continue to provide good genetic care for the inhabitants of the geneti-cally isolated population under study and to continue to adjust the care to the demands of the population.
Currently, several aspects and developments are important to take into consideration. • Until recently, the majority of inhabitants of the genetically isolated population
under study knew someone with one of the four (five) severe recessive disorders. However, after the setup of the outpatient clinic, presumably the number of children born with one of these disorders has been reduced drastically (personal communica-tion). Long term follow up is needed to assess the actual impact of screening. Parallel to this reduction, the awareness of the severity and impact of these disorders will presumably diminish. It will be a challenge to keep the inhabitants fully informed of the disorders.
• The question is whether ancestry-based carrier screening is the preferred way of carrier screening in this and other genetically isolated populations, or whether we should offer population-based ECS, in which a wide array of diseases is screened not aimed solely at specific high-risk groups. The five severe recessive disorders frequently found in the genetically isolated population are included in the population-based ECS panel in the Netherlands and this panel is thus suitable for use in this population. We suspect that the mutations in other genes included in this ECS panel are not present in the isolated population because no patients with these disorders are known in the
9
population. However, as population-based ECS might decrease possible stigmatiza-tion by offering a ‘universal’ test rather than targeted to a particular subpopulastigmatiza-tion, this way of carrier screening should be considered,52 taking into account the costs of the test(s) and the demands of the population.
• We experience that genetic counseling in the genetically isolated population is becoming increasingly complex, as illustrated by the pedigree in Figure 1. Besides the four (five) severe recessive disorders included in the carrier screening panel, other (frequently) less-severe recessive disorders are also highly frequent. We offer addi-tional genetic counseling for these disorders if (one of these) disorders are present in the family or if the counselees ask for further information about these disorders. The question is whether carrier screening for these disorders has to be introduced in a more systematic way by offering one or more separate test panels (see under Carrier screening in genetically isolated populations).
• During pregnancy, cell-free fetal (placental) DNA (cfDNA) is circulating freely in maternal blood. This cfDNA can be used for non-invasive prenatal testing (NIPT) to screening for fetal aneuploidy. In the Netherlands, NIPT was first offered only to women at increased risk of fetal aneuploidy,78,79 but is since April 2017 avail-able to all pregnant women within the TRIDENT-2 study. The discovery of cfDNA also enables non-invasive prenatal diagnosis for monogenic disorders (MG-NIPD).
Figure 1. Example of a pedigree of a carrier couple for OI and PCD identified by carrier screening in the
genetically isolated population under study. The woman is affected with PKU.
Squares indicate males; circles, females; diamonds, gender unspecified; arrows, probands; shaded symbols, affected; spot, carrier; PKU, phenylketonuria; OI, osteogenesis imperfecta type IIB/III; FADS, fetal akinesia deformation sequence; PCD, primary ciliary dyskinesia; POLG, Polymerase gamma (POLG)-related disorders; IUFD, intrauterine fetal death.
NIPD for de novo80 and paternally inherited autosomal dominant conditions81 is relatively straightforward and is already offered for some conditions. However, NIPD for autosomal recessive and de novo maternal autosomal dominant condi-tions remains challenging because of the predominance of the maternal mutant allele in the cfDNA sample.82 The first proof-of-concept studies have been performed.83 NIPD for the five severe autosomal recessive disorders in the genetically isolated population under study may even be more complicated because the quantity of informative SNPs may be reduced as a consequence of the common ancestry of the couples. The demands of the carrier couples for developing a NIPD test for these disorders are high and we are currently trying to set up NIPD for these disorders. • Knowing which disorders are more prevalent in genetically isolated populations, including
their specific genes and mutations, is important for making a rapid (differential) diag-nosis by clinicians, genetic counseling, and research in these populations. Currently, such a database is not available in the Netherlands but we are considering setting up a database. It is still debated whether this database should be an open access database or, to prevent stigmatization, a database only accessible for health care providers.
CONCLUSION
Genetically isolated populations provide unique opportunities for many different genetic investigations. This is illustrated in this thesis by different investigations in a young genetically isolated population in the Netherlands. In this population we identified two new founder mutations, we studied genetic heterogeneity, phenotypic variation, and we implemented and evaluated carrier screening for multiple recessive disorders to enable reproductive choices. Although genetically isolated populations may differ in various aspects from other populations and not all research findings may be representative for the general population, investigations in these populations may be helpful in under-standing causes of genetic disorders and may provide lessons for further successful and responsible implementation of expanded carrier screening.
9
REFERENCES
1. Lander ES, Botstein D. Homozygosity mapping: a way to map human recessive traits with the DNA of inbred children. Science. 1987;236:1567-1570.
2. Tan-Sindhunata MB, Mathijssen IB, Smit M, Baas F, de Vries JI, van der Voorn JP, Kluijt I, Hagen MA, Blom EW, Sistermans E, Meijers-Heijboer H, Waisfisz Q, Weiss MM, Groffen AJ. Identification of a Dutch founder mutation in MUSK causing fetal akinesia deformation sequence. Eur J Hum Genet. 2015;23:1151-1157.
3. Heutink P, Oostra BA. Gene finding in genetically isolated populations. Hum Mol Genet. 2002;11:2507-2515. 4. Zeggini E. Using genetically isolated populations to understand the genomic basis of disease. Genome Med. 2014;6:83. 5. Onoufriadis A, Paff T, Antony D, Shoemark A, Micha D, Kuyt B, Schmidts M, Petridi S, Dankert-Roelse JE,
Haarman EG, Daniels JM, Emes RD, Wilson R, Hogg C, Scambler PJ, Chung EM, Pals G, Mitchison HM. Splice-site mutations in the axonemal outer dynein arm docking complex gene CCDC114 cause primary ciliary dyskinesia. Am J Hum Genet. 2013;92:88-98.
6. Van Dijk FS, Nesbitt IM, Nikkels PG, Dalton A, Bongers EM, van de Kamp JM, Hilhorst-Hofstee Y, den Hollander NS, Lachmeijer AM, Marcelis CL, Tan-Sindhunata GM, van Rijn RR, Meijers-Heijboer H, Cobben JM, Pals G. CRTAP mutations in lethal and severe osteogenesis imperfecta: the importance of combining biochemical and molecular genetic analysis. Eur J Hum Genet. 2009;17:1560-1569.
7. Wilbe M, Ekvall S, Eurenius K, Ericson K, Casar-Borota O, Klar J, Dahl N, Ameur A, Anneren G, Bondeson ML. MuSK: a new target for lethal fetal akinesia deformation sequence (FADS). J Med Genet. 2015;52:195-202. 8. Chatzispyrou IA, Alders M, Guerrero-Castillo S, Zapata PR, Haagmans MA, Mouchiroud L, Koster J, Ofman R, Baas F,
Waterham HR, Spelbrink JN, Auwerx J, Mannens MM, Houtkooper RH, Plomp AS. A homozygous missense mutation in ERAL1, encoding a mitochondrial rRNA chaperone, causes Perrault syndrome. Hum Mol Genet. 2017;26:2541-2550. 9. Bleeker-Wagemakers LM, Gal A, Kumar-Singh R, van den Born LI, Li Y, Schwinger E, Sandkuijl LA, Bergen AA,
Kenna P, Humphries P. Evidence for nonallelic genetic heterogeneity in autosomal recessive retinitis pigmentosa. Genomics. 1992;14:811-812.
10. Van den Born LI, Bergen AA, Bleeker-Wagemakers EM. A retrospective study of registered retinitis pigmentosa patients in The Netherlands. Ophthalmic Paediatr Genet. 1992;13:227-236.
11. Van den Born LI, van Soest S, van Schooneveld MJ, Riemslag FC, de Jong PT, Bleeker-Wagemakers EM. Auto-somal recessive retinitis pigmentosa with preserved para-arteriolar retinal pigment epithelium. Am J Ophthalmol. 1994;118:430-439.
12. Van Soest S, van den Born LI, Gal A, Farrar GJ, Bleeker-Wagemakers LM, Westerveld A, Humphries P, Sandkuijl LA, Bergen AA. Assignment of a gene for autosomal recessive retinitis pigmentosa (RP12) to chromosome 1q31-q32.1 in an inbred and genetically heterogeneous disease population. Genomics. 1994;22:499-504.
13. Den Hollander AI, ten Brink JB, de Kok YJ, van Soest S, van den Born LI, van Driel MA, van de Pol DJ, Payne AM, Bhattacharya SS, Kellner U, Hoyng CB, Westerveld A, Brunner HG, Bleeker-Wagemakers EM, Deutman AF, Heckenlively JR, Cremers FP, Bergen AA. Mutations in a human homologue of Drosophila crumbs cause retinitis pigmentosa (RP12). Nat Genet. 1999;23:217-221.
14. Mathijssen IB, Florijn RJ, van den Born LI, Zekveld-Vroon RC, ten Brink JB, Plomp AS, Baas F, Meijers-Heijboer H, Bergen AA, van Schooneveld MJ. Long-term follow-up of patients with retinitis pigmentosa type 12 caused by CRB1 mutations: A severe phenotype with considerable interindividual variability. Retina. 2017;37:161-172. 15. Adato A, Raskin L, Petit C, Bonne-Tamir B. Deafness heterogeneity in a Druze isolate from the Middle East: novel
OTOF and PDS mutations, low prevalence of GJB2 35delG mutation and indication for a new DFNB locus. Eur J Hum Genet. 2000;8:437-442.
16. Carrasquillo MM, Zlotogora J, Barges S, Chakravarti A. Two different connexin 26 mutations in an inbred kindred segregating non-syndromic recessive deafness: implications for genetic studies in isolated populations. Hum Mol Genet. 1997;6:2163-2172.
17. Plomp AS, Bergen AA, Florijn RJ, Terry SF, Toonstra J, van Dijk MR, de Jong PT. Pseudoxanthoma elasticum: Wide phenotypic variation in homozygotes and no signs in heterozygotes for the c.3775delT mutation in ABCC6. Genet Med. 2009;11:852-858.
18. Mathijssen IB, Henneman L, van Eeten-Nijman JM, Lakeman P, Ottenheim CP, Redeker EJ, Ottenhof W, Meijers-Heijboer H, van Maarle MC. Targeted carrier screening for four recessive disorders: high detection rate within a founder population. Eur J Med Genet. 2015;58:123-128.
19. Oorthuys JW, Gons MH, Kwakman JV, Schutgens RB, Tegelaers WH. [Screening for raised serum phenylalanine levels in women of reproductive age in Volendam]. Ned Tijdschr Geneeskd. 1985;129:62-65.
20. Barth PG, Vrensen GF, Uylings HB, Oorthuys JW, Stam FC. Inherited syndrome of microcephaly, dyskinesia and pontocerebellar hypoplasia: a systemic atrophy with early onset. J Neurol Sci. 1990;97:25-42.
21. Barth PG, Blennow G, Lenard HG, Begeer JH, van der Kley JM, Hanefeld F, Peters AC, Valk J. The syndrome of autosomal recessive pontocerebellar hypoplasia, microcephaly, and extrapyramidal dyskinesia (pontocerebellar hypoplasia type 2): compiled data from 10 pedigrees. Neurology. 1995;45:311-317.
22. Barth PG, Aronica E, de Vries L, Nikkels PG, Scheper W, Hoozemans JJ, BT P-T, Troost D. Pontocerebellar hypo-plasia type 2: a neuropathological update. Acta Neuropathol. 2007;114:373-386.
23. Budde BS, Namavar Y, Barth PG, BT P-T, Nurnberg G, Becker C, van Ruissen F, Weterman MA, Fluiter K, te Beek ET, Aronica E, van der Knaap MS, Hohne W, Toliat MR, Crow YJ, Steinling M, Voit T, Roelenso F, Brussel W, Brockmann K, Kyllerman M, Boltshauser E, Hammersen G, Willemsen M, Basel-Vanagaite L, Krageloh-Mann I, de Vries LS, Sztriha L, Muntoni F, Ferrie CD, Battini R, Hennekam RC, Grillo E, Beemer FA, Stoets LM, Wollnik B, Nurnberg P, Baas F. tRNA splicing endonuclease mutations cause pontocerebellar hypoplasia. Nat Genet. 2008;40:1113-1118.
24. Bergen AA, Plomp AS, Schuurman EJ, Terry S, Breuning M, Dauwerse H, Swart J, Kool M, van Soest S, Baas F, ten Brink JB, de Jong PT. Mutations in ABCC6 cause pseudoxanthoma elasticum. Nat Genet. 2000;25:228-231. 25. Van Soest S, Swart J, Tijmes N, Sandkuijl LA, Rommers J, Bergen AA. A locus for autosomal recessive
pseudoxan-thoma elasticum, with penetrance of vascular symptoms in carriers, maps to chromosome 16p13.1. Genome Res. 1997;7:830-834.
26. Handley MT, Morris-Rosendahl DJ, Brown S, Macdonald F, Hardy C, Bem D, Carpanini SM, Borck G, Martorell L, Izzi C, Faravelli F, Accorsi P, Pinelli L, Basel-Vanagaite L, Peretz G, Abdel-Salam GM, Zaki MS, Jansen A, Mowat D, Glass I, Stewart H, Mancini G, Lederer D, Roscioli T, Giuliano F, Plomp AS, Rolfs A, Graham JM, Seemanova E, Poo P, Garcia-Cazorla A, Edery P, Jackson IJ, Maher ER, Aligianis IA. Mutation spectrum in RAB3GAP1, RAB3GAP2, and RAB18 and genotype-phenotype correlations in warburg micro syndrome and Martsolf syndrome. Hum Mutat. 2013;34:686-696.
27. Mathijssen IB, van Maarle MC, Kleiss IIM, Redeker EJW, Ten Kate LP, Henneman L, Meijers-Heijboer H. With expanded carrier screening, founder populations run the risk of being overlooked. J Community Genet. 2017;8:327-333. 28. Zeegers MP, van Poppel F, Vlietinck R, Spruijt L, Ostrer H. Founder mutations among the Dutch. Eur J Hum
Genet. 2004;12:591-600.
29. Resta RG. Defining and redefining the scope and goals of genetic counseling. Am J Med Genet C Semin Med Genet. 2006;142C:269-275.
30. Payne M, Rupar CA, Siu GM, Siu VM. Amish, mennonite, and hutterite genetic disorder database. Paediatr Child Health. 2011;16:e23-e24.
31. Polvi A, Linturi H, Varilo T, Anttonen AK, Byrne M, Fokkema IF, Almusa H, Metzidis A, Avela K, Aula P, Kestila M, Muilu J. The Finnish disease heritage database (FinDis) update-a database for the genes mutated in the Finnish disease heritage brought to the next-generation sequencing era. Hum Mutat. 2013;34:1458-1466.
32. Zlotogora J. The molecular basis of autosomal recessive diseases among the Arabs and Druze in Israel. Hum Genet. 2010;128:473-479.
9
33. Genetics ACo. ACOG Committee Opinion No. 442: Preconception and prenatal carrier screening for genetic diseases in individuals of Eastern European Jewish descent. Obstet Gynecol. 2009;114:950-953.
34. Basel-Vanagaite L, Taub E, Halpern GJ, Drasinover V, Magal N, Davidov B, Zlotogora J, Shohat M. Genetic screening for autosomal recessive nonsyndromic mental retardation in an isolated population in Israel. Eur J Hum Genet. 2007;15:250-253.
35. Falik-Zaccai TC, Kfir N, Frenkel P, Cohen C, Tanus M, Mandel H, Shihab S, Morkos S, Aaref S, Summar ML, Khayat M. Population screening in a Druze community: the challenge and the reward. Genet Med. 2008;10:903-909. 36. Zlotogora J, Carmi R, Lev B, Shalev SA. A targeted population carrier screening program for severe and frequent
genetic diseases in Israel. Eur J Hum Genet. 2009;17:591-597.
37. Tardif J, Pratte A, Laberge AM. Experience of carrier couples identified through a population-based carrier screening pilot program for four founder autosomal recessive diseases in Saguenay-Lac-Saint-Jean. Prenat Diagn. 2017;doi: 10.1002/pd.5055.
38. Holtkamp KCA, Mathijssen IB, Lakeman P, van Maarle MC, Dondorp WJ, Henneman L, Cornel MC. Factors for successful implementation of population-based expanded carrier screening: learning from existing initiatives. Eur J Public Health. 2017;27:372-377.
39. Borry P, Henneman L, Lakeman P, ten Kate LP, Cornel MC, Howard HC. Preconceptional genetic carrier testing and the commercial offer directly-to-consumers. Hum Reprod. 2011;26:972-977.
40. Grody WW, Thompson BH, Gregg AR, Bean LH, Monaghan KG, Schneider A, Lebo RV. ACMG position state-ment on prenatal/preconception expanded carrier screening. Genet Med. 2013;15:482-483.
41. Borry P, Clarke A, Dierickx K. Look before you leap. Carrier screening for type 1 Gaucher disease: difficult ques-tions. Eur J Hum Genet. 2008;16:139-140.
42. Beutler E. Carrier screening for Gaucher disease: more harm than good? JAMA. 2007;298:1329-1331.
43. Nielsen M, Morreau H, Vasen HF, Hes FJ. MUTYH-associated polyposis (MAP). Crit Rev Oncol Hematol. 2011;79:1-16.
44. Al-Tassan N, Chmiel NH, Maynard J, Fleming N, Livingston AL, Williams GT, Hodges AK, Davies DR, David SS, Sampson JR. Inherited variants of MYH associated with somatic G: C [arrow right] T: A mutations in colorectal tumors. Nature genetics. 2002;30:227.
45. Croitoru ME, Cleary SP, Di Nicola N, Manno M, Selander T, Aronson M, Redston M, Cotterchio M, Knight J, Gryfe R. Association between biallelic and monoallelic germline MYH gene mutations and colorectal cancer risk. Journal of the National Cancer Institute. 2004;96:1631-1634.
46. Struewing JP, Hartge P, Wacholder S, Baker SM, Berlin M, McAdams M, Timmerman MM, Brody LC, Tucker MA. The risk of cancer associated with specific mutations of BRCA1 and BRCA2 among Ashkenazi Jews. N Engl J Med. 1997;336:1401-1408.
47. Gabai-Kapara E, Lahad A, Kaufman B, Friedman E, Segev S, Renbaum P, Beeri R, Gal M, Grinshpun-Cohen J, Djemal K, Mandell JB, Lee MK, Beller U, Catane R, King MC, Levy-Lahad E. Population-based screening for breast and ovarian cancer risk due to BRCA1 and BRCA2. Proc Natl Acad Sci U S A. 2014;111:14205-14210. 48. Manchanda R, Loggenberg K, Sanderson S, Burnell M, Wardle J, Gessler S, Side L, Balogun N, Desai R, Kumar
A, Dorkins H, Wallis Y, Chapman C, Taylor R, Jacobs C, Tomlinson I, McGuire A, Beller U, Menon U, Jacobs I. Population testing for cancer predisposing BRCA1/BRCA2 mutations in the Ashkenazi-Jewish community: a rand-omized controlled trial. J Natl Cancer Inst. 2015;107:379.
49. Metcalfe KA, Poll A, Royer R, Llacuachaqui M, Tulman A, Sun P, Narod SA. Screening for founder mutations in BRCA1 and BRCA2 in unselected Jewish women. J Clin Oncol. 2010;28:387-391.
50. Metcalfe KA, Eisen A, Lerner-Ellis J, Narod SA. Is it time to offer BRCA1 and BRCA2 testing to all Jewish women? Curr Oncol. 2015;22:e233-236.
51. Edwards JG, Feldman G, Goldberg J, Gregg AR, Norton ME, Rose NC, Schneider A, Stoll K, Wapner R, Watson MS. Expanded carrier screening in reproductive medicine-points to consider: a joint statement of the American College of Medical Genetics and Genomics, American College of Obstetricians and Gynecologists, National Society of Genetic Counselors, Perinatal Quality Foundation, and Society for Maternal-Fetal Medicine. Obstet Gynecol. 2015;125:653-662.
52. Henneman L, Borry P, Chokoshvili D, Cornel MC, van El CG, Forzano F, Hall A, Howard HC, Janssens S, Kayserili H, Lakeman P, Lucassen A, Metcalfe SA, Vidmar L, de Wert G, Dondorp WJ, Peterlin B. Responsible implementation of expanded carrier screening. Eur J Hum Genet. 2016;24:e1-e12.
53. Lazarin GA, Haque IS, Nazareth S, Iori K, Patterson AS, Jacobson JL, Marshall JR, Seltzer WK, Patrizio P, Evans EA, Srinivasan BS. An empirical estimate of carrier frequencies for 400+ causal Mendelian variants: results from an ethnically diverse clinical sample of 23,453 individuals. Genet Med. 2013;15:178-186.
54. Holtkamp KC, van Maarle MC, Schouten MJ, Dondorp WJ, Lakeman P, Henneman L. Do people from the Jewish community prefer ancestry-based or pan-ethnic expanded carrier screening? Eur J Hum Genet. 2016;24:171-177. 55. Sallevelt SC, de Koning B, Szklarczyk R, Paulussen AD, de Die-Smulders CE, Smeets HJ. A comprehensive strategy
for exome-based preconception carrier screening. Genet Med. 2017;19:583-592.
56. Teeuw M, Waisfisz Q, Zwijnenburg PJ, Sistermans EA, Weiss MM, Henneman L, ten Kate LP, Cornel MC, Meijers-Heijboer H. First steps in exploring prospective exome sequencing of consanguineous couples. Eur J Med Genet. 2014;57:613-616.
57. Beaudet AL. Global genetic carrier testing: a vision for the future. Genome Med. 2015;7:79.
58. Ioannou L, Massie J, Lewis S, Petrou V, Gason A, Metcalfe S, Aitken MA, Bankier A, Delatycki MB. Evaluation of a multi-disease carrier screening programme in Ashkenazi Jewish high schools. Clin Genet. 2010;78:21-31. 59. Rothwell E, Johnson E, Mathiesen A, Golden K, Metcalf A, Rose NC, Botkin JR. Experiences among Women with
Positive Prenatal Expanded Carrier Screening Results. J Genet Couns. 2017;26:690-696.
60. Warsch JR, Warsch S, Herman E, Zakarin L, Schneider A, Hoffman J, Wasserman D, Barbouth D. Knowledge, atti-tudes, and barriers to carrier screening for the Ashkenazi Jewish panel: a Florida experience : Education and Barriers assessment for Jewish Genetic Diseases. J Community Genet. 2014;5:223-231.
61. Cho D, McGowan ML, Metcalfe J, Sharp RR. Expanded carrier screening in reproductive healthcare: perspectives from genetics professionals. Hum Reprod. 2013;28:1725-1730.
62. Plantinga M, Birnie E, Abbott KM, Sinke RJ, Lucassen AM, Schuurmans J, Kaplan S, Verkerk MA, Ranchor AV, van Langen IM. Population-based preconception carrier screening: how potential users from the general population view a test for 50 serious diseases. Eur J Hum Genet. 2016;24:1417-1423.
63. Fan CW, Castonguay L, Rummell S, Levesque S, Mitchell JJ, Sillon G. Online Module for Carrier Screening in Ashkenazi Jewish Individuals Compared with In-Person Genetics Education: A Randomized Controlled Trial. J Genet Couns. 2017;doi:10.1007/s10897-017-0133-4.
64. Brandt-Rauf SI, Raveis VH, Drummond NF, Conte JA, Rothman SM. Ashkenazi Jews and breast cancer: the conse-quences of linking ethnic identity to genetic disease. Am J Public Health. 2006;96:1979-1988.
65. Lazarin GA, Haque IS. Expanded carrier screening: A review of early implementation and literature. Semin Peri-natol. 2016;40:29-34.
66. Van der Hout S, Holtkamp KC, Henneman L, de Wert G, Dondorp WJ. Advantages of expanded universal carrier screening: what is at stake? Eur J Hum Genet. 2016;25:17-21.
67. Raz AE, Vizner Y. Carrier matching and collective socialization in community genetics: Dor Yeshorim and the rein-forcement of stigma. Soc Sci Med. 2008;67:1361-1369.
68. Gordon C, Walpole I, Zubrick SR, Bower C. Population screening for cystic fibrosis: knowledge and emotional consequences 18 months later. Am J Med Genet A. 2003;120A:199-208.
9
69. Leib JR, Gollust SE, Hull SC, Wilfond BS. Carrier screening panels for Ashkenazi Jews: is more better? Genet Med. 2005;7:185-190.
70. Harper PS, Clarke AJ. Genetics Society and Clinical Practice. Oxford, UK: BIOS Scientific Publishers, 1997. 71. De Wert GM, Dondorp WJ, Knoppers BM. Preconception care and genetic risk: ethical issues. J Community
Genet. 2012;3:221-228.
72. Godard B, ten Kate L, Evers-Kiebooms G, Ayme S. Population genetic screening programmes: principles, tech-niques, practices, and policies. Eur J Hum Genet. 2003;11 Suppl 2:S49-87.
73. Jackson L, Goldsmith L, O’Connor A, Skirton H. Incidental findings in genetic research and clinical diagnostic tests: a systematic review. Am J Med Genet A. 2012;158A:3159-3167.
74. Lemke AA, Wolf WA, Hebert-Beirne J, Smith ME. Public and biobank participant attitudes toward genetic research participation and data sharing. Public Health Genomics. 2010;13:368-377.
75. Kristiansson K, Naukkarinen J, Peltonen L. Isolated populations and complex disease gene identification. Genome Biol. 2008;9:109.
76. Peltonen L, Palotie A, Lange K. Use of population isolates for mapping complex traits. Nat Rev Genet. 2000;1:182-190.
77. Takahashi K, Yamanaka S. Induction of pluripotent stem cells from mouse embryonic and adult fibroblast cultures by defined factors. Cell. 2006;126:663-676.
78. Oepkes D, Yaron Y, Kozlowski P, Rego de Sousa MJ, Bartha JL, van den Akker ES, Dornan SM, Krampl-Bettelheim E, Schmid M, Wielgos M, Cirigliano V, Di Renzo GC, Cameron A, Calda P, Tabor A. Counseling for non-invasive prenatal testing (NIPT): what pregnant women may want to know. Ultrasound Obstet Gynecol. 2014;44:1-5. 79. Van Schendel RV, Page-Christiaens GC, Beulen L, Bilardo CM, de Boer MA, Coumans AB, Faas BH, van Langen
IM, Lichtenbelt KD, van Maarle MC, Macville MV, Oepkes D, Pajkrt E, Henneman L, Dutch NC. Trial by Dutch laboratories for evaluation of non-invasive prenatal testing. Part II-women’s perspectives. Prenat Diagn. 2016;36:1091-1098.
80. Chitty LS, Griffin DR, Meaney C, Barrett A, Khalil A, Pajkrt E, Cole TJ. New aids for the non-invasive prenatal diagnosis of achondroplasia: dysmorphic features, charts of fetal size and molecular confirmation using cell-free fetal DNA in maternal plasma. Ultrasound Obstet Gynecol. 2011;37:283-289.
81. Van den Oever JM, Bijlsma EK, Feenstra I, Muntjewerff N, Mathijssen IB, Bakker E, van Belzen MJ, Boon EM. Noninvasive prenatal diagnosis of Huntington disease: detection of the paternally inherited expanded CAG repeat in maternal plasma. Prenat Diagn. 2015;35:945-949.
82. Drury S, Mason S, McKay F, Lo K, Boustred C, Jenkins L, Chitty LS. Implementing Non-Invasive Prenatal Diag-nosis (NIPD) in a National Health Service Laboratory; From Dominant to Recessive Disorders. Adv Exp Med Biol. 2016;924:71-75.
83. Vermeulen C, Geeven G, de Wit E, Verstegen M, Jansen RPM, van Kranenburg M, de Bruijn E, Pulit SL, Kruis-selbrink E, Shahsavari Z, Omrani D, Zeinali F, Najmabadi H, Katsila T, Vrettou C, Patrinos GP, Traeger-Synodinos J, Splinter E, Beekman JM, Kheradmand Kia S, Te Meerman GJ, Ploos van Amstel HK, de Laat W. Sensitive Mono-genic Noninvasive Prenatal Diagnosis by Targeted Haplotyping. Am J Hum Genet. 2017;101:326-339.
